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Method DNA extraction
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DNA EXTRACTION
Grade 10 Science Life and Living DNA
Compiled by Madrersquo Nortje
2
SCIENCE WORKEXPERIMENT 21
DNA EXTRACTION
A CONTEXTUAL APPROACH SCIENCE 10 (HEINEMANN) P125
GRADE 10 Life and Living
DATE of Practical 17 March 2011
3
QUESTION WHAT DOES DNA STAND FOR
DEOXYRIBONUCLEIC ACID
THE COMPLEX CHEMICAL COMPOUND FOUND INN CHROMOSOMES THAT CONTAINS THE GENETIC CODE
TODAY WE WILL CONNECT DNA WITHextraction
DO ONLY LIVING ORGANISMS CONTAIN DNA
What do you think
Discuss in pairs for 1 minute
Give reasons
5
EXTRACTING DNA FROM ANY LIVING THING
Just follow these 3 easy steps
DetergenteNzymes Alcohol
httplearngeneticsutaheducontentlabsextractionhowto
6
YO
U M
EA
N I
CA
N S
EE I
T
HO
W
DNA -
Deoxyribonucle
ic acid
bullAll living organisms contain DNA in their cells ndash FOUND in nucleus
bullDNA - The complex chemical compound found in chromosomes that
contains the genetic code
The nucleus is a membrane bound structure that contains the cells hereditary information and controls the cells growth and reproduction It is commonly the most prominent organelle in the cell
wwwkamibudaksainsblogspotcom
7
FIGURE HTTPEMPLOYEESCSBSJUEDUHJAKUBOWSKICLASSESCH331DNAOLDNASTRUCTUREHTML
PACKAGING OF DNA IN THE NUCLEUS
8
httpwwwyoutubecomwatchv=tmVP1cotozc
9
DNA extraction refers to the process that scientists use to break down a piece of evidence such as a piece of hair or a drop of blood in order to determine the individuals DNA which is unique to each person Understand the process of DNA extraction with information from a biology teacher in this free video on science
Expert Janice CrenettiContact WeAreHDTVcomBio Janice Creneti has a Bachelor of Science in secondary science education and a Bachelor of Art in biology from Boston UniversityFilmmaker Christopher Rokosz
Category Entertainment Tags science dna dna testing dna structure dna model dna replication dna molecule
10
REVIEWING OF INFORMATION ONLABORATORY SAFETY RULES AND RISK ASSESSMENTS
Do not enter the laboratory unless you are with a teacher Never touch equipment in the laboratory unless you are told to use it Donrsquot eat or drink in the laboratory Always walkmdashnever run Wear protective clothingmdash a laboratory coat or apron and when
appropriate safety glasses Never taste anything Tie up long hair Always point test tubes away from people Check with your teacher on how to dispose of waste liquids and solids
Broken glass should be cleaned up using gloves a brush and dustpan and placed in a special bin
If you spill something on your skin or clothes wash it immediately with lots of water Tell your teacher
Report all accidents and breakages to your teacher After heating equipment let it cool on a heatproof mat before picking
it up This will avoid burns Clean all equipment after use and put it back where you got it from
Clean and dry your work bench
11
RISK ASSESSMENT ON EXPERIMENT AND INVESTIGATION ndash KIWI FRUIT
Eyewear and protective clothingNo eat or drinkingCover the MSDSrsquo e on Isopropanol and Methylene blue ndash hard copy handouts (health and safety regulation)
Any broken glassware must be reported
Care must be taken when using and washing the blender ndash sharp blades
12
ISOPROPANOL ndash MSDS HAND OUTS CHEMWATCH ISSUE DATE 15052010 ISOPROPANOL - FLAMMABLE Dangerous Good 3 SAFETY-DATA(tm) Ratings (Provided here for your convenience)
----------------------------------------------------------------------------------------------------------- Health Rating 2 - Moderate Flammability Rating 3 - Severe (Flammable) Reactivity Rating 2 - Moderate Contact Rating 3 - Severe Lab Protective Equip GOGGLES amp SHIELD LAB COAT amp APRON VENT HOOD PROPER GLOVES CLASS B EXTINGUISHER Storage Colour Code Red (Flammable)
Potential Health Effects ----------------------------------
Inhalation Inhalation of vapours irritates the respiratory tract Exposure to high concentrations has a narcotic effect producing symptoms of dizziness drowsiness headache staggering unconsciousness and possibly death Ingestion Can cause drowsiness unconsciousness and death Gastrointestinal pain cramps nausea vomiting and diarrhoea may also result The single lethal dose for a human adult = about 250 mls (8 ounces) Skin Contact May cause irritation with redness and pain May be absorbed through the skin with possible systemic effects Eye Contact Vapours cause eye irritation Splashes cause severe irritation possible corneal burns and eye damage Chronic Exposure Chronic exposure may cause skin effects Aggravation of Pre-existing Conditions Persons with pre-existing skin disorders or impaired liver kidney or pulmonary function may be more susceptible to the effects of this agent
13
METHYLENE BLUE ndash MSDS HAND OUTS CHEMWATCH ISSUE DATE 15052010 METHYLENE BLUE SAFETY-DATA(tm) Ratings (Provided here for your convenience)
----------------------------------------------------------------------------------------------------------- Health Rating 2 - Moderate Flammability Rating 1 - Slight Reactivity Rating 1 - Slight Contact Rating 1 - Slight Lab Protective Equip GOGGLES LAB COAT VENT HOOD PROPER GLOVES Storage Colour Code Green (General Storage) -----------------------------------------------------------------------------------------------------------
Potential Health Effects ----------------------------------
This material is relatively nonhazardous in routine industrial situations
Inhalation No adverse health effects expected from inhalation May cause a short period of rapid or difficult breathing Ingestion A burning sensation of the mouth may be noted following ingestion of methylene blue May cause nausea vomiting diarrhea and gastritis Large doses may cause abdominal and chest pain headache profuse sweating mental confusion painful maturation and methemoglobinemia Skin Contact Not expected to be a health hazard from skin exposure Methylene blue may colour the skin a bluish colour May cause photosensitization Eye Contact No adverse effects expected May cause mechanical irritation Chronic Exposure No information found Aggravation of Pre-existing Conditions No information found
14
NEW WORDS TO WORD BANKADD THESE TO YOUR EXCEL ALPHA TABLE
Precipitate Spooling Denaturising Lysing Homogenized Aggregate Lipid Centrifuging PCR
WORDS MEANING
15
bullDO YOU THINK YOU HAVE VERY MUCH IN COMMON WITH A KIWI FRUIT
BELIEVE IT OR NOT A KIWIS GENETIC MATERIAL IS VERY SIMILAR TO YOUR OWN
SEE AND TOUCH THE GENETIC MATERIAL THAT YOULL EXTRACT FROM THE CELLS OF A KIWI FRUIT
Activity Overview
Extract DNA from kiwi fruit using simple household chemicals
16
HT
TP
LEA
RN
GEN
ETIC
SU
TA
HE
DU
CO
NTE
NTL
AB
SE
XTR
AC
TIO
NH
OW
TO
DNA EXTRACTION USING KIWI FRUIT
17
AIM
TO
EX
TR
AC
T D
NA
FR
OM
TH
E C
ELLS
OF K
IWI F
RU
IT
wwwexploratoriumedu
MATERIALS EQUIPMENT (per student group takes 30 minutes)
frac12 cup Kiwi fruit 200 mL water dishwashing detergent dropping pipette fine mesh kitchen strainer amp cheese cloth glass rod large beaker large test tube light microscope meat tenderiser methylene blue microscope lamp inoculation needle microscope slide and cover slip paper towelling small beaker of alcohol (Isopropanol) spatula test-tube rack mortar and pestle
18
PART A EXTRACTING THE DNA
METHOD
1 Peal and Place the kiwi and water in the beaker and mush it with a mortar
and pestle DNA SOURCE About 125 ml 12 cup Twice as much water as DNA source (250
ml 1 cup) Table salt large pinch 1g 14 teaspoon Stir until the mixture is of a thin soupy
consistency
TAKE NOTES ndash NO TEXTBOOKS ALLOWED WHEN PERFORMING EXPERIMENT ndashMUST RECALL PROCEDURES METHOD
19
WHAT MUST I DO NOW
STRAIN THE DNA MIXTURE
DISHWASHING DETERGENT
20
2 Pour the thin cell mixture through the kitchen strainer (cheesecloth) into another large beaker
3 Measure 12 ml of the soup into
a small beakerAdd 2 ml of liquid detergentSwirl to mix stir thoroughly using a
glass rodLet mixture sit in container with
hot tap water (60 - 65degC) for 5 - 10 minutes
21
WHAT MUST I DO NOWMEAT TENDERIZER
ENZYME POWDERALCOHOL SEPARATION
wwwforumsoverclockerscomau
22
4 Add a pinch of enzyme (meat tenderizer) to mixture
(Using pineapple juice or contact lens cleaning solution will do the same as tenderizer)
Gently stir with toothpick skewer for 5 minutes continue stirring not too vigorously Be careful If you stir too hard youll break up the DNA making it harder to see
5 Quarter-fill a large test tube with the mixture
23
SLOWLY pour the same amount ice - cold (70 - 95 isopropyl or ethyl alcohol) into test tube pour alcohol down the side of the test tube
Tilting the test tube will make this easier to do
It forms a layer on top of the cell mixture
DO NOT MIX THE TWO LAYERS TOGETHER
Amount of alcohol and mixture should be the same
24
SEPARATION USING ALCOHOL
25
7 Observe the mixture for a few minutes You will see a white threadlike substance rise from the mixture to rest above in the alcohol layer
This is the DNA that you have extracted from the cells of the kiwi fruit
8 You can get more DNA to precipitate from the solution using a DNA collecting tool (glass or paper clip hook or cut inoculation needle)
Gently lift the water solution up into the alcohol layer (this allows more DNA to get in contact with the alcohol and precipitate)
26
INFORMATION DNA precipitates as a white stringy
ldquosnottyrdquo film at the water - alcohol interface and eventually will rise into the alcohol layer from the mixture layer
Allow test tube to sit for several minutes The clearer the DNA is the fewer
impurities you have
If you have an acceptable amount of DNA it can be spooled by rotating your collecting tool and then transferred into a clean tube container
27
WOW ndash SPOOLING THE DNA If you are careful you may be able wind up the DNA
around a glass rod or a skewer Position the tip of the glass rod or skewer where you can see the threads of DNA Steadily twist the rod or skewer as if you were making candy floss Alternatively use a straw to pull it out by suction ndash be careful not to get in you mouth
Donrsquot go too quickly
You should be able to pull the strands of DNA out of the mixture
ASK STUDENTS TO TAKE MOBILE PHONE PICTURES OF THEIR OWN EXTRACTED DNA AND COMPARE APPEARANCE WITH OTHERS
Photos of steps can be inserted in flow diagram(ICT)
28
KIWIDNA
29
If you want to save your DNA you can transfer it to a small container filled with alcohol
Leave tube container uncapped until the ethanol has evaporated
DNA can be stored in the fridge (dry or water buffer can be added)
You can now investigate the property
30
WHAT IS THAT STRINGY STUFFDNA IS A LONG STRINGY MOLECULE THE SALT THAT YOU ADDED IN STEP ONE HELPS IT STICK TOGETHER SO WHAT YOU SEE ARE CLUMPS OF TANGLED DNA MOLECULES
DNA NORMALLY STAYS DISSOLVED IN WATER BUT WHEN SALTY DNA COMES IN CONTACT WITH ALCOHOL IT BECOMES UNDISSOLVED THIS IS CALLED PRECIPITATION THE PHYSICAL FORCE OF THE DNA CLUMPING TOGETHER AS IT PRECIPITATES PULLS MORE STRANDS ALONG WITH IT AS IT RISES INTO THE ALCOHOL
31
THE WHYrsquoS Blending separated the pea cells In order to extract DNA from a cell the associated
membranes and proteins must first be removed (break apart the cells) and then physically separated (loosen the tough cell wall) from the DNA
To see the DNA we have to break open these two sacks We do this with detergent and salt( Sodium can be
involved in several of the steps ) Sodium is an element Its chemical symbol is Na for
Natrium the Latin word for sodium It is a positive ion and often associates with negative ions as part of useful compounds
Salt help precipitate protein and carbohydrates away from the DNA
Salt helps strip away the proteins associated with DNA + and ndash charge
32
THE WHYrsquoS Salt and Detergents are used to break down
cell walls and nuclear membranes to release the DNA
They work by chemically poking holes in the cell membranes or walls
Once holes are poked in the membranes the membranes can be further disrupted mechanically as with a blender
After that it is easier to get the contents of the cell out including the DNA
LETS HAVE A LOOK AT THE CELL
33
CELL TO DNA
httplearngeneticsutaheducontentlabsextractionhowtoHOW TO EXTRACT DNA FROM ANYTHING LIVING
34
THE WHYrsquoS WHY DETERGENT
Each cell is surrounded by a sack (the cell membrane)
DNA is found inside the second sack (nucleus) within each cell
To see the DNA we have to break it open A cell membrane has 2 layers of lipid(fat)
molecules with protein going through them When the lysis buffer (detergent) comes
close to the cell it captures the lipids and the proteins - breaks open the cell destroying the fatty membrane - DNA now released into the solution
35
A CELLS MEMBRANES HAVE TWO LAYERS OF LIPID (FAT) MOLECULES WITH PROTEINS GOING THROUGH THEM
36
Why detergent How does detergent work
Think about why you use soap to wash dishes or your hands To remove grease and dirt right
Soap molecules and grease molecules are made of two parts
1(Blue) Heads which like water 2(Green) Tails which hate water
37
AFTER ADDING THE DETERGENT WHAT DO YOU HAVE IN YOUR SOUP
38
THE WHYrsquoS ENZYME (MEAT TENDERIZER)
The DNA in the nucleus of the cell is molded folded and protected by proteins The tenderizer cut the proteins away from the DNA
In this experiment meat tenderizer acts as an enzyme to cut proteins just like a pair of scissors
The meat tenderizer cuts the proteins away from the DNA
39
THE WHYrsquoS CUTTING PROTEIN AND DNA
The DNA in the nucleus of the cell is moulded folded and protected by proteins
40
THE WHYrsquoSALCOHOL Alcohol is less dense than water so it floats
on top Look for clumps of white stringy stuff where the water and alcohol layers meet
DNA is not soluble in alcohol - other cell parts are
By adding alcohol DNA precipitates out of the solution and collect at the interface of the alcohol and soap layer
The colder the alcohol the less soluble the DNA will be in it
41
COLD ALCOHOL
DNA dissolves in water but precipitates in alcohol
Cold alcohol is used to separate DNA out of water-based solutions
This allows the DNA to be purified for subsequent genetic testing
Adding alcohol to a solution containing DNA is a simple way to obtain the pure DNA required and colder temperatures slow down enzymes that can break down DNA giving better extraction results
42
WHY IS DNA EXTRACTION IMPORTANT DNA extraction is an important molecular
biology procedure
By definition extraction is taking DNA out of any type of cell for the purpose of analysis
See Handouts for more information on the use of DNA extracted (extension only)
43
SUMMARIZEDNA EXTRACTING FROM KIWI FRUIT
44
BREAK DOWN CELL WALLS
Cells walls have to be destroyed to reach the DNA within cells
Extraction procedures to obtain pure DNA have to get rid of all the molecular and chemical components of the tissue from which the DNA is being extracted
First steps involve lysing or destroying the cell walls This can be done with a variety of chemical agents that are caustic to cell membranes but do not harm the DNA Cells can also be sonicated homogenized or ground up to destroy membranes
45
REMOVING Removing Lipids
Once the cell walls have been destroyed a detergent is added to get rid of the fats and oils that make up the cell membranes Detergents cause the fats and oils to dissolve into the solution
Remove ProteinsProteins and enzymes can be digested by
adding a protease to the solution Proteases break down proteins into small peptides and amino acids
46
PRECIPITATING AND PURIFYING Precipitate DNA
Adding cold alcohol to a solution will cause DNA to precipitate and aggregate The DNA can be collected by centrifuging the sample and pouring off the liquid layer The DNA should exist as a small pellet in the bottom of the centrifuge tube
Purify DNAThe DNA can be washed by re-suspending it
in a cold alcohol solution and re-centrifuging it several times to obtain a very pure DNA sample Typical alcohols used to precipitate DNA include ethanol and Isopropanol This process leaves a very pure sample for stringent DNA testing
47
PART B OBSERVING EXTRACTED DNA UNDER THE MICROSCOPE
48
PART B A CLOSER LOOK
8 Use a dropping pipette to carefully remove some of the threadlike substance from the top of your preparation
9 Place one or two drops onto the middle of a
microscope slide
10 Add two drops of methylene blue Wait 3 or 4 minutes to allow the methylene blue to be absorbed by the DNA
11 Carefully place a cover slip on the slide Gently press
a folded piece of paper towelling over the top of the prepared slide to soak up any excess liquid
12 Observe the DNA under low power then high power
49
DISCUSSIONANSWER IN GROUP RELATED WORKED 1 Write a detailed description of the material
floating at the top of the test tube after the alcohol was added
2 Describe the DNA as it appears under the
microscope under high power 3 Prepare a diagram of your DNA specimen 4 What was the reason for using methylene
blue
50
HOMEWORKCONSTRUCT A FLOW CHART DRAWING MIND MAP OF THE PROCESS YOU USED EXTRACTING THE DNA FROM KIWIrsquoS
Next to each step explain how the method you used was important (Hint What substances make up the membranes of cells and cell organelles
How do detergents work
What is the active ingredient in meat tenderiser
51
TIPS STEPS FOR FLOW CHART In order to release the DNA from the
nuclei of the pea cells you first separated the cells from one another
Then the cell and nuclear membranes needed to be ruptured to release the cell contents and the contents of the nucleus
Once removed from the nuclear membrane the DNA had to be untangled into the visible threadlike structures you ended up with
52
HOMEWORK EXTENSION GROUP QUESTIONS Where can DNA be found in the cell Discuss the action of the soap (detergent) on the cell
What is the purpose of the soap in this activity What was the purpose of the Sodium Chloride Include a
discussion of polarity and charged particles Why was the cold ethanol added to the soap and salt
mixture Describe the appearance of your final product Draw a diagram of DNA containing 5 sets of nucleotide
bases labelling the hydrogen bonds between the bases References and Resources Adapted from Berry Full
of DNA by Diane Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
53
HTTPWWWGENOMEBCCAEDUCATIONTEACHERSCLASSROOM-ACTIVITIESDNA-EXTRACTION
54
EXTRACHROMOSOMES EXIST IN PAIRS Because our chromosomes exist in pairs (and consequently we
have 2 alleles of each gene) we are a diploid species This is why our somatic cells are represented as 2n Our gametes (sperm and ova) on the other hand are haploid and are represented as n Other species may have different ploidy for example
triploid (3n) seedless watermelons tetaploid (4n) salmonidae fish pentaploid (5n) Kenai birch hexaploid (6n) some types of wheat kiwi fruit octaploid (8n) acipenser (a genus
of sturgeon fish strawberies) decaploid (10n) some strawberries dodecaploid (12n) some types of amphibians eg Xenopus
ruwenzoriensis
wwwcyberusca
55
YOUR EXTRACTED DNA UNDER THE MICROSCOPE LOOKED
LIKE
Collaboration-Brainstorming on what to look at under the
- DNA extraction- DNA is too small for even a microscope to see and after the extraction it just appears like a blob True or false
Could you any DNA strand
wwwemployeescsbsjuedu
56
HANDOUT ON DNA EXTRACTIONBACKGROUND EVERY DAY LIFE
EXTRA READING MATERIAL(EXTENSION )
DISCOVERING DNA DNA ON THE INSIDE USE OF DNA EXTRACTION INN EVERYDAY LIFE WHY IS DNA TESTING GOOD MOLECULAR BIOLOGY PCR-based diagnostics of genomic DNA
57
REFERENCES httpwwwsquidoocomhow-to-get-dna-from-a-kiwi-fruit Why Is DNA Testing Good | eHowcom
httpwwwehowcomabout_6684410_dna-testing-good_htmlixzz1MkbcIJs5 Why Is DNA Extraction Important | eHowcom
httpwwwehowcomlist_5839095_dna-extraction-important_htmlixzz1MkZDrqyY
Why Is Sodium Used in DNA Extraction | eHowcom httpwwwehowcomabout_6504902_sodium-used-dna-extraction_htmlixzz1MkaGWg57
Why Is Cold Alcohol Used in DNA Tests | eHowcom httpwwwehowcomabout_6399349_cold-alcohol-used-dna-tests_htmlixzz1MkbEmEdu
httplearngeneticsutaheducontentlabsextractionhowto www kamibudaksainsblogspotcom
httpwwwchemwatch goldcom References and Resources Adapted from Berry Full of DNA by Diane
Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
58
REFERENCES
Uses of DNA Extraction | eHowcom httpwwwehowcomabout_5344428_uses-dna-extractionhtmlixzz1MkWQB2TZ
Science 10 A Contextual Approach Heineman Queensland Science Projects Regan Spence Maggie Spenceley
httpenwikipediaorgwikiMolecular_biology httpwwwclinchemorgcgicontent
extract44102201 http
employeescsbsjueduhjakubowskiclassesch331dnaoldnastructurehtml
httpwwwgenomebccaeducationteachersclassroom-activitiesdna-extraction
2
SCIENCE WORKEXPERIMENT 21
DNA EXTRACTION
A CONTEXTUAL APPROACH SCIENCE 10 (HEINEMANN) P125
GRADE 10 Life and Living
DATE of Practical 17 March 2011
3
QUESTION WHAT DOES DNA STAND FOR
DEOXYRIBONUCLEIC ACID
THE COMPLEX CHEMICAL COMPOUND FOUND INN CHROMOSOMES THAT CONTAINS THE GENETIC CODE
TODAY WE WILL CONNECT DNA WITHextraction
DO ONLY LIVING ORGANISMS CONTAIN DNA
What do you think
Discuss in pairs for 1 minute
Give reasons
5
EXTRACTING DNA FROM ANY LIVING THING
Just follow these 3 easy steps
DetergenteNzymes Alcohol
httplearngeneticsutaheducontentlabsextractionhowto
6
YO
U M
EA
N I
CA
N S
EE I
T
HO
W
DNA -
Deoxyribonucle
ic acid
bullAll living organisms contain DNA in their cells ndash FOUND in nucleus
bullDNA - The complex chemical compound found in chromosomes that
contains the genetic code
The nucleus is a membrane bound structure that contains the cells hereditary information and controls the cells growth and reproduction It is commonly the most prominent organelle in the cell
wwwkamibudaksainsblogspotcom
7
FIGURE HTTPEMPLOYEESCSBSJUEDUHJAKUBOWSKICLASSESCH331DNAOLDNASTRUCTUREHTML
PACKAGING OF DNA IN THE NUCLEUS
8
httpwwwyoutubecomwatchv=tmVP1cotozc
9
DNA extraction refers to the process that scientists use to break down a piece of evidence such as a piece of hair or a drop of blood in order to determine the individuals DNA which is unique to each person Understand the process of DNA extraction with information from a biology teacher in this free video on science
Expert Janice CrenettiContact WeAreHDTVcomBio Janice Creneti has a Bachelor of Science in secondary science education and a Bachelor of Art in biology from Boston UniversityFilmmaker Christopher Rokosz
Category Entertainment Tags science dna dna testing dna structure dna model dna replication dna molecule
10
REVIEWING OF INFORMATION ONLABORATORY SAFETY RULES AND RISK ASSESSMENTS
Do not enter the laboratory unless you are with a teacher Never touch equipment in the laboratory unless you are told to use it Donrsquot eat or drink in the laboratory Always walkmdashnever run Wear protective clothingmdash a laboratory coat or apron and when
appropriate safety glasses Never taste anything Tie up long hair Always point test tubes away from people Check with your teacher on how to dispose of waste liquids and solids
Broken glass should be cleaned up using gloves a brush and dustpan and placed in a special bin
If you spill something on your skin or clothes wash it immediately with lots of water Tell your teacher
Report all accidents and breakages to your teacher After heating equipment let it cool on a heatproof mat before picking
it up This will avoid burns Clean all equipment after use and put it back where you got it from
Clean and dry your work bench
11
RISK ASSESSMENT ON EXPERIMENT AND INVESTIGATION ndash KIWI FRUIT
Eyewear and protective clothingNo eat or drinkingCover the MSDSrsquo e on Isopropanol and Methylene blue ndash hard copy handouts (health and safety regulation)
Any broken glassware must be reported
Care must be taken when using and washing the blender ndash sharp blades
12
ISOPROPANOL ndash MSDS HAND OUTS CHEMWATCH ISSUE DATE 15052010 ISOPROPANOL - FLAMMABLE Dangerous Good 3 SAFETY-DATA(tm) Ratings (Provided here for your convenience)
----------------------------------------------------------------------------------------------------------- Health Rating 2 - Moderate Flammability Rating 3 - Severe (Flammable) Reactivity Rating 2 - Moderate Contact Rating 3 - Severe Lab Protective Equip GOGGLES amp SHIELD LAB COAT amp APRON VENT HOOD PROPER GLOVES CLASS B EXTINGUISHER Storage Colour Code Red (Flammable)
Potential Health Effects ----------------------------------
Inhalation Inhalation of vapours irritates the respiratory tract Exposure to high concentrations has a narcotic effect producing symptoms of dizziness drowsiness headache staggering unconsciousness and possibly death Ingestion Can cause drowsiness unconsciousness and death Gastrointestinal pain cramps nausea vomiting and diarrhoea may also result The single lethal dose for a human adult = about 250 mls (8 ounces) Skin Contact May cause irritation with redness and pain May be absorbed through the skin with possible systemic effects Eye Contact Vapours cause eye irritation Splashes cause severe irritation possible corneal burns and eye damage Chronic Exposure Chronic exposure may cause skin effects Aggravation of Pre-existing Conditions Persons with pre-existing skin disorders or impaired liver kidney or pulmonary function may be more susceptible to the effects of this agent
13
METHYLENE BLUE ndash MSDS HAND OUTS CHEMWATCH ISSUE DATE 15052010 METHYLENE BLUE SAFETY-DATA(tm) Ratings (Provided here for your convenience)
----------------------------------------------------------------------------------------------------------- Health Rating 2 - Moderate Flammability Rating 1 - Slight Reactivity Rating 1 - Slight Contact Rating 1 - Slight Lab Protective Equip GOGGLES LAB COAT VENT HOOD PROPER GLOVES Storage Colour Code Green (General Storage) -----------------------------------------------------------------------------------------------------------
Potential Health Effects ----------------------------------
This material is relatively nonhazardous in routine industrial situations
Inhalation No adverse health effects expected from inhalation May cause a short period of rapid or difficult breathing Ingestion A burning sensation of the mouth may be noted following ingestion of methylene blue May cause nausea vomiting diarrhea and gastritis Large doses may cause abdominal and chest pain headache profuse sweating mental confusion painful maturation and methemoglobinemia Skin Contact Not expected to be a health hazard from skin exposure Methylene blue may colour the skin a bluish colour May cause photosensitization Eye Contact No adverse effects expected May cause mechanical irritation Chronic Exposure No information found Aggravation of Pre-existing Conditions No information found
14
NEW WORDS TO WORD BANKADD THESE TO YOUR EXCEL ALPHA TABLE
Precipitate Spooling Denaturising Lysing Homogenized Aggregate Lipid Centrifuging PCR
WORDS MEANING
15
bullDO YOU THINK YOU HAVE VERY MUCH IN COMMON WITH A KIWI FRUIT
BELIEVE IT OR NOT A KIWIS GENETIC MATERIAL IS VERY SIMILAR TO YOUR OWN
SEE AND TOUCH THE GENETIC MATERIAL THAT YOULL EXTRACT FROM THE CELLS OF A KIWI FRUIT
Activity Overview
Extract DNA from kiwi fruit using simple household chemicals
16
HT
TP
LEA
RN
GEN
ETIC
SU
TA
HE
DU
CO
NTE
NTL
AB
SE
XTR
AC
TIO
NH
OW
TO
DNA EXTRACTION USING KIWI FRUIT
17
AIM
TO
EX
TR
AC
T D
NA
FR
OM
TH
E C
ELLS
OF K
IWI F
RU
IT
wwwexploratoriumedu
MATERIALS EQUIPMENT (per student group takes 30 minutes)
frac12 cup Kiwi fruit 200 mL water dishwashing detergent dropping pipette fine mesh kitchen strainer amp cheese cloth glass rod large beaker large test tube light microscope meat tenderiser methylene blue microscope lamp inoculation needle microscope slide and cover slip paper towelling small beaker of alcohol (Isopropanol) spatula test-tube rack mortar and pestle
18
PART A EXTRACTING THE DNA
METHOD
1 Peal and Place the kiwi and water in the beaker and mush it with a mortar
and pestle DNA SOURCE About 125 ml 12 cup Twice as much water as DNA source (250
ml 1 cup) Table salt large pinch 1g 14 teaspoon Stir until the mixture is of a thin soupy
consistency
TAKE NOTES ndash NO TEXTBOOKS ALLOWED WHEN PERFORMING EXPERIMENT ndashMUST RECALL PROCEDURES METHOD
19
WHAT MUST I DO NOW
STRAIN THE DNA MIXTURE
DISHWASHING DETERGENT
20
2 Pour the thin cell mixture through the kitchen strainer (cheesecloth) into another large beaker
3 Measure 12 ml of the soup into
a small beakerAdd 2 ml of liquid detergentSwirl to mix stir thoroughly using a
glass rodLet mixture sit in container with
hot tap water (60 - 65degC) for 5 - 10 minutes
21
WHAT MUST I DO NOWMEAT TENDERIZER
ENZYME POWDERALCOHOL SEPARATION
wwwforumsoverclockerscomau
22
4 Add a pinch of enzyme (meat tenderizer) to mixture
(Using pineapple juice or contact lens cleaning solution will do the same as tenderizer)
Gently stir with toothpick skewer for 5 minutes continue stirring not too vigorously Be careful If you stir too hard youll break up the DNA making it harder to see
5 Quarter-fill a large test tube with the mixture
23
SLOWLY pour the same amount ice - cold (70 - 95 isopropyl or ethyl alcohol) into test tube pour alcohol down the side of the test tube
Tilting the test tube will make this easier to do
It forms a layer on top of the cell mixture
DO NOT MIX THE TWO LAYERS TOGETHER
Amount of alcohol and mixture should be the same
24
SEPARATION USING ALCOHOL
25
7 Observe the mixture for a few minutes You will see a white threadlike substance rise from the mixture to rest above in the alcohol layer
This is the DNA that you have extracted from the cells of the kiwi fruit
8 You can get more DNA to precipitate from the solution using a DNA collecting tool (glass or paper clip hook or cut inoculation needle)
Gently lift the water solution up into the alcohol layer (this allows more DNA to get in contact with the alcohol and precipitate)
26
INFORMATION DNA precipitates as a white stringy
ldquosnottyrdquo film at the water - alcohol interface and eventually will rise into the alcohol layer from the mixture layer
Allow test tube to sit for several minutes The clearer the DNA is the fewer
impurities you have
If you have an acceptable amount of DNA it can be spooled by rotating your collecting tool and then transferred into a clean tube container
27
WOW ndash SPOOLING THE DNA If you are careful you may be able wind up the DNA
around a glass rod or a skewer Position the tip of the glass rod or skewer where you can see the threads of DNA Steadily twist the rod or skewer as if you were making candy floss Alternatively use a straw to pull it out by suction ndash be careful not to get in you mouth
Donrsquot go too quickly
You should be able to pull the strands of DNA out of the mixture
ASK STUDENTS TO TAKE MOBILE PHONE PICTURES OF THEIR OWN EXTRACTED DNA AND COMPARE APPEARANCE WITH OTHERS
Photos of steps can be inserted in flow diagram(ICT)
28
KIWIDNA
29
If you want to save your DNA you can transfer it to a small container filled with alcohol
Leave tube container uncapped until the ethanol has evaporated
DNA can be stored in the fridge (dry or water buffer can be added)
You can now investigate the property
30
WHAT IS THAT STRINGY STUFFDNA IS A LONG STRINGY MOLECULE THE SALT THAT YOU ADDED IN STEP ONE HELPS IT STICK TOGETHER SO WHAT YOU SEE ARE CLUMPS OF TANGLED DNA MOLECULES
DNA NORMALLY STAYS DISSOLVED IN WATER BUT WHEN SALTY DNA COMES IN CONTACT WITH ALCOHOL IT BECOMES UNDISSOLVED THIS IS CALLED PRECIPITATION THE PHYSICAL FORCE OF THE DNA CLUMPING TOGETHER AS IT PRECIPITATES PULLS MORE STRANDS ALONG WITH IT AS IT RISES INTO THE ALCOHOL
31
THE WHYrsquoS Blending separated the pea cells In order to extract DNA from a cell the associated
membranes and proteins must first be removed (break apart the cells) and then physically separated (loosen the tough cell wall) from the DNA
To see the DNA we have to break open these two sacks We do this with detergent and salt( Sodium can be
involved in several of the steps ) Sodium is an element Its chemical symbol is Na for
Natrium the Latin word for sodium It is a positive ion and often associates with negative ions as part of useful compounds
Salt help precipitate protein and carbohydrates away from the DNA
Salt helps strip away the proteins associated with DNA + and ndash charge
32
THE WHYrsquoS Salt and Detergents are used to break down
cell walls and nuclear membranes to release the DNA
They work by chemically poking holes in the cell membranes or walls
Once holes are poked in the membranes the membranes can be further disrupted mechanically as with a blender
After that it is easier to get the contents of the cell out including the DNA
LETS HAVE A LOOK AT THE CELL
33
CELL TO DNA
httplearngeneticsutaheducontentlabsextractionhowtoHOW TO EXTRACT DNA FROM ANYTHING LIVING
34
THE WHYrsquoS WHY DETERGENT
Each cell is surrounded by a sack (the cell membrane)
DNA is found inside the second sack (nucleus) within each cell
To see the DNA we have to break it open A cell membrane has 2 layers of lipid(fat)
molecules with protein going through them When the lysis buffer (detergent) comes
close to the cell it captures the lipids and the proteins - breaks open the cell destroying the fatty membrane - DNA now released into the solution
35
A CELLS MEMBRANES HAVE TWO LAYERS OF LIPID (FAT) MOLECULES WITH PROTEINS GOING THROUGH THEM
36
Why detergent How does detergent work
Think about why you use soap to wash dishes or your hands To remove grease and dirt right
Soap molecules and grease molecules are made of two parts
1(Blue) Heads which like water 2(Green) Tails which hate water
37
AFTER ADDING THE DETERGENT WHAT DO YOU HAVE IN YOUR SOUP
38
THE WHYrsquoS ENZYME (MEAT TENDERIZER)
The DNA in the nucleus of the cell is molded folded and protected by proteins The tenderizer cut the proteins away from the DNA
In this experiment meat tenderizer acts as an enzyme to cut proteins just like a pair of scissors
The meat tenderizer cuts the proteins away from the DNA
39
THE WHYrsquoS CUTTING PROTEIN AND DNA
The DNA in the nucleus of the cell is moulded folded and protected by proteins
40
THE WHYrsquoSALCOHOL Alcohol is less dense than water so it floats
on top Look for clumps of white stringy stuff where the water and alcohol layers meet
DNA is not soluble in alcohol - other cell parts are
By adding alcohol DNA precipitates out of the solution and collect at the interface of the alcohol and soap layer
The colder the alcohol the less soluble the DNA will be in it
41
COLD ALCOHOL
DNA dissolves in water but precipitates in alcohol
Cold alcohol is used to separate DNA out of water-based solutions
This allows the DNA to be purified for subsequent genetic testing
Adding alcohol to a solution containing DNA is a simple way to obtain the pure DNA required and colder temperatures slow down enzymes that can break down DNA giving better extraction results
42
WHY IS DNA EXTRACTION IMPORTANT DNA extraction is an important molecular
biology procedure
By definition extraction is taking DNA out of any type of cell for the purpose of analysis
See Handouts for more information on the use of DNA extracted (extension only)
43
SUMMARIZEDNA EXTRACTING FROM KIWI FRUIT
44
BREAK DOWN CELL WALLS
Cells walls have to be destroyed to reach the DNA within cells
Extraction procedures to obtain pure DNA have to get rid of all the molecular and chemical components of the tissue from which the DNA is being extracted
First steps involve lysing or destroying the cell walls This can be done with a variety of chemical agents that are caustic to cell membranes but do not harm the DNA Cells can also be sonicated homogenized or ground up to destroy membranes
45
REMOVING Removing Lipids
Once the cell walls have been destroyed a detergent is added to get rid of the fats and oils that make up the cell membranes Detergents cause the fats and oils to dissolve into the solution
Remove ProteinsProteins and enzymes can be digested by
adding a protease to the solution Proteases break down proteins into small peptides and amino acids
46
PRECIPITATING AND PURIFYING Precipitate DNA
Adding cold alcohol to a solution will cause DNA to precipitate and aggregate The DNA can be collected by centrifuging the sample and pouring off the liquid layer The DNA should exist as a small pellet in the bottom of the centrifuge tube
Purify DNAThe DNA can be washed by re-suspending it
in a cold alcohol solution and re-centrifuging it several times to obtain a very pure DNA sample Typical alcohols used to precipitate DNA include ethanol and Isopropanol This process leaves a very pure sample for stringent DNA testing
47
PART B OBSERVING EXTRACTED DNA UNDER THE MICROSCOPE
48
PART B A CLOSER LOOK
8 Use a dropping pipette to carefully remove some of the threadlike substance from the top of your preparation
9 Place one or two drops onto the middle of a
microscope slide
10 Add two drops of methylene blue Wait 3 or 4 minutes to allow the methylene blue to be absorbed by the DNA
11 Carefully place a cover slip on the slide Gently press
a folded piece of paper towelling over the top of the prepared slide to soak up any excess liquid
12 Observe the DNA under low power then high power
49
DISCUSSIONANSWER IN GROUP RELATED WORKED 1 Write a detailed description of the material
floating at the top of the test tube after the alcohol was added
2 Describe the DNA as it appears under the
microscope under high power 3 Prepare a diagram of your DNA specimen 4 What was the reason for using methylene
blue
50
HOMEWORKCONSTRUCT A FLOW CHART DRAWING MIND MAP OF THE PROCESS YOU USED EXTRACTING THE DNA FROM KIWIrsquoS
Next to each step explain how the method you used was important (Hint What substances make up the membranes of cells and cell organelles
How do detergents work
What is the active ingredient in meat tenderiser
51
TIPS STEPS FOR FLOW CHART In order to release the DNA from the
nuclei of the pea cells you first separated the cells from one another
Then the cell and nuclear membranes needed to be ruptured to release the cell contents and the contents of the nucleus
Once removed from the nuclear membrane the DNA had to be untangled into the visible threadlike structures you ended up with
52
HOMEWORK EXTENSION GROUP QUESTIONS Where can DNA be found in the cell Discuss the action of the soap (detergent) on the cell
What is the purpose of the soap in this activity What was the purpose of the Sodium Chloride Include a
discussion of polarity and charged particles Why was the cold ethanol added to the soap and salt
mixture Describe the appearance of your final product Draw a diagram of DNA containing 5 sets of nucleotide
bases labelling the hydrogen bonds between the bases References and Resources Adapted from Berry Full
of DNA by Diane Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
53
HTTPWWWGENOMEBCCAEDUCATIONTEACHERSCLASSROOM-ACTIVITIESDNA-EXTRACTION
54
EXTRACHROMOSOMES EXIST IN PAIRS Because our chromosomes exist in pairs (and consequently we
have 2 alleles of each gene) we are a diploid species This is why our somatic cells are represented as 2n Our gametes (sperm and ova) on the other hand are haploid and are represented as n Other species may have different ploidy for example
triploid (3n) seedless watermelons tetaploid (4n) salmonidae fish pentaploid (5n) Kenai birch hexaploid (6n) some types of wheat kiwi fruit octaploid (8n) acipenser (a genus
of sturgeon fish strawberies) decaploid (10n) some strawberries dodecaploid (12n) some types of amphibians eg Xenopus
ruwenzoriensis
wwwcyberusca
55
YOUR EXTRACTED DNA UNDER THE MICROSCOPE LOOKED
LIKE
Collaboration-Brainstorming on what to look at under the
- DNA extraction- DNA is too small for even a microscope to see and after the extraction it just appears like a blob True or false
Could you any DNA strand
wwwemployeescsbsjuedu
56
HANDOUT ON DNA EXTRACTIONBACKGROUND EVERY DAY LIFE
EXTRA READING MATERIAL(EXTENSION )
DISCOVERING DNA DNA ON THE INSIDE USE OF DNA EXTRACTION INN EVERYDAY LIFE WHY IS DNA TESTING GOOD MOLECULAR BIOLOGY PCR-based diagnostics of genomic DNA
57
REFERENCES httpwwwsquidoocomhow-to-get-dna-from-a-kiwi-fruit Why Is DNA Testing Good | eHowcom
httpwwwehowcomabout_6684410_dna-testing-good_htmlixzz1MkbcIJs5 Why Is DNA Extraction Important | eHowcom
httpwwwehowcomlist_5839095_dna-extraction-important_htmlixzz1MkZDrqyY
Why Is Sodium Used in DNA Extraction | eHowcom httpwwwehowcomabout_6504902_sodium-used-dna-extraction_htmlixzz1MkaGWg57
Why Is Cold Alcohol Used in DNA Tests | eHowcom httpwwwehowcomabout_6399349_cold-alcohol-used-dna-tests_htmlixzz1MkbEmEdu
httplearngeneticsutaheducontentlabsextractionhowto www kamibudaksainsblogspotcom
httpwwwchemwatch goldcom References and Resources Adapted from Berry Full of DNA by Diane
Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
58
REFERENCES
Uses of DNA Extraction | eHowcom httpwwwehowcomabout_5344428_uses-dna-extractionhtmlixzz1MkWQB2TZ
Science 10 A Contextual Approach Heineman Queensland Science Projects Regan Spence Maggie Spenceley
httpenwikipediaorgwikiMolecular_biology httpwwwclinchemorgcgicontent
extract44102201 http
employeescsbsjueduhjakubowskiclassesch331dnaoldnastructurehtml
httpwwwgenomebccaeducationteachersclassroom-activitiesdna-extraction
3
QUESTION WHAT DOES DNA STAND FOR
DEOXYRIBONUCLEIC ACID
THE COMPLEX CHEMICAL COMPOUND FOUND INN CHROMOSOMES THAT CONTAINS THE GENETIC CODE
TODAY WE WILL CONNECT DNA WITHextraction
DO ONLY LIVING ORGANISMS CONTAIN DNA
What do you think
Discuss in pairs for 1 minute
Give reasons
5
EXTRACTING DNA FROM ANY LIVING THING
Just follow these 3 easy steps
DetergenteNzymes Alcohol
httplearngeneticsutaheducontentlabsextractionhowto
6
YO
U M
EA
N I
CA
N S
EE I
T
HO
W
DNA -
Deoxyribonucle
ic acid
bullAll living organisms contain DNA in their cells ndash FOUND in nucleus
bullDNA - The complex chemical compound found in chromosomes that
contains the genetic code
The nucleus is a membrane bound structure that contains the cells hereditary information and controls the cells growth and reproduction It is commonly the most prominent organelle in the cell
wwwkamibudaksainsblogspotcom
7
FIGURE HTTPEMPLOYEESCSBSJUEDUHJAKUBOWSKICLASSESCH331DNAOLDNASTRUCTUREHTML
PACKAGING OF DNA IN THE NUCLEUS
8
httpwwwyoutubecomwatchv=tmVP1cotozc
9
DNA extraction refers to the process that scientists use to break down a piece of evidence such as a piece of hair or a drop of blood in order to determine the individuals DNA which is unique to each person Understand the process of DNA extraction with information from a biology teacher in this free video on science
Expert Janice CrenettiContact WeAreHDTVcomBio Janice Creneti has a Bachelor of Science in secondary science education and a Bachelor of Art in biology from Boston UniversityFilmmaker Christopher Rokosz
Category Entertainment Tags science dna dna testing dna structure dna model dna replication dna molecule
10
REVIEWING OF INFORMATION ONLABORATORY SAFETY RULES AND RISK ASSESSMENTS
Do not enter the laboratory unless you are with a teacher Never touch equipment in the laboratory unless you are told to use it Donrsquot eat or drink in the laboratory Always walkmdashnever run Wear protective clothingmdash a laboratory coat or apron and when
appropriate safety glasses Never taste anything Tie up long hair Always point test tubes away from people Check with your teacher on how to dispose of waste liquids and solids
Broken glass should be cleaned up using gloves a brush and dustpan and placed in a special bin
If you spill something on your skin or clothes wash it immediately with lots of water Tell your teacher
Report all accidents and breakages to your teacher After heating equipment let it cool on a heatproof mat before picking
it up This will avoid burns Clean all equipment after use and put it back where you got it from
Clean and dry your work bench
11
RISK ASSESSMENT ON EXPERIMENT AND INVESTIGATION ndash KIWI FRUIT
Eyewear and protective clothingNo eat or drinkingCover the MSDSrsquo e on Isopropanol and Methylene blue ndash hard copy handouts (health and safety regulation)
Any broken glassware must be reported
Care must be taken when using and washing the blender ndash sharp blades
12
ISOPROPANOL ndash MSDS HAND OUTS CHEMWATCH ISSUE DATE 15052010 ISOPROPANOL - FLAMMABLE Dangerous Good 3 SAFETY-DATA(tm) Ratings (Provided here for your convenience)
----------------------------------------------------------------------------------------------------------- Health Rating 2 - Moderate Flammability Rating 3 - Severe (Flammable) Reactivity Rating 2 - Moderate Contact Rating 3 - Severe Lab Protective Equip GOGGLES amp SHIELD LAB COAT amp APRON VENT HOOD PROPER GLOVES CLASS B EXTINGUISHER Storage Colour Code Red (Flammable)
Potential Health Effects ----------------------------------
Inhalation Inhalation of vapours irritates the respiratory tract Exposure to high concentrations has a narcotic effect producing symptoms of dizziness drowsiness headache staggering unconsciousness and possibly death Ingestion Can cause drowsiness unconsciousness and death Gastrointestinal pain cramps nausea vomiting and diarrhoea may also result The single lethal dose for a human adult = about 250 mls (8 ounces) Skin Contact May cause irritation with redness and pain May be absorbed through the skin with possible systemic effects Eye Contact Vapours cause eye irritation Splashes cause severe irritation possible corneal burns and eye damage Chronic Exposure Chronic exposure may cause skin effects Aggravation of Pre-existing Conditions Persons with pre-existing skin disorders or impaired liver kidney or pulmonary function may be more susceptible to the effects of this agent
13
METHYLENE BLUE ndash MSDS HAND OUTS CHEMWATCH ISSUE DATE 15052010 METHYLENE BLUE SAFETY-DATA(tm) Ratings (Provided here for your convenience)
----------------------------------------------------------------------------------------------------------- Health Rating 2 - Moderate Flammability Rating 1 - Slight Reactivity Rating 1 - Slight Contact Rating 1 - Slight Lab Protective Equip GOGGLES LAB COAT VENT HOOD PROPER GLOVES Storage Colour Code Green (General Storage) -----------------------------------------------------------------------------------------------------------
Potential Health Effects ----------------------------------
This material is relatively nonhazardous in routine industrial situations
Inhalation No adverse health effects expected from inhalation May cause a short period of rapid or difficult breathing Ingestion A burning sensation of the mouth may be noted following ingestion of methylene blue May cause nausea vomiting diarrhea and gastritis Large doses may cause abdominal and chest pain headache profuse sweating mental confusion painful maturation and methemoglobinemia Skin Contact Not expected to be a health hazard from skin exposure Methylene blue may colour the skin a bluish colour May cause photosensitization Eye Contact No adverse effects expected May cause mechanical irritation Chronic Exposure No information found Aggravation of Pre-existing Conditions No information found
14
NEW WORDS TO WORD BANKADD THESE TO YOUR EXCEL ALPHA TABLE
Precipitate Spooling Denaturising Lysing Homogenized Aggregate Lipid Centrifuging PCR
WORDS MEANING
15
bullDO YOU THINK YOU HAVE VERY MUCH IN COMMON WITH A KIWI FRUIT
BELIEVE IT OR NOT A KIWIS GENETIC MATERIAL IS VERY SIMILAR TO YOUR OWN
SEE AND TOUCH THE GENETIC MATERIAL THAT YOULL EXTRACT FROM THE CELLS OF A KIWI FRUIT
Activity Overview
Extract DNA from kiwi fruit using simple household chemicals
16
HT
TP
LEA
RN
GEN
ETIC
SU
TA
HE
DU
CO
NTE
NTL
AB
SE
XTR
AC
TIO
NH
OW
TO
DNA EXTRACTION USING KIWI FRUIT
17
AIM
TO
EX
TR
AC
T D
NA
FR
OM
TH
E C
ELLS
OF K
IWI F
RU
IT
wwwexploratoriumedu
MATERIALS EQUIPMENT (per student group takes 30 minutes)
frac12 cup Kiwi fruit 200 mL water dishwashing detergent dropping pipette fine mesh kitchen strainer amp cheese cloth glass rod large beaker large test tube light microscope meat tenderiser methylene blue microscope lamp inoculation needle microscope slide and cover slip paper towelling small beaker of alcohol (Isopropanol) spatula test-tube rack mortar and pestle
18
PART A EXTRACTING THE DNA
METHOD
1 Peal and Place the kiwi and water in the beaker and mush it with a mortar
and pestle DNA SOURCE About 125 ml 12 cup Twice as much water as DNA source (250
ml 1 cup) Table salt large pinch 1g 14 teaspoon Stir until the mixture is of a thin soupy
consistency
TAKE NOTES ndash NO TEXTBOOKS ALLOWED WHEN PERFORMING EXPERIMENT ndashMUST RECALL PROCEDURES METHOD
19
WHAT MUST I DO NOW
STRAIN THE DNA MIXTURE
DISHWASHING DETERGENT
20
2 Pour the thin cell mixture through the kitchen strainer (cheesecloth) into another large beaker
3 Measure 12 ml of the soup into
a small beakerAdd 2 ml of liquid detergentSwirl to mix stir thoroughly using a
glass rodLet mixture sit in container with
hot tap water (60 - 65degC) for 5 - 10 minutes
21
WHAT MUST I DO NOWMEAT TENDERIZER
ENZYME POWDERALCOHOL SEPARATION
wwwforumsoverclockerscomau
22
4 Add a pinch of enzyme (meat tenderizer) to mixture
(Using pineapple juice or contact lens cleaning solution will do the same as tenderizer)
Gently stir with toothpick skewer for 5 minutes continue stirring not too vigorously Be careful If you stir too hard youll break up the DNA making it harder to see
5 Quarter-fill a large test tube with the mixture
23
SLOWLY pour the same amount ice - cold (70 - 95 isopropyl or ethyl alcohol) into test tube pour alcohol down the side of the test tube
Tilting the test tube will make this easier to do
It forms a layer on top of the cell mixture
DO NOT MIX THE TWO LAYERS TOGETHER
Amount of alcohol and mixture should be the same
24
SEPARATION USING ALCOHOL
25
7 Observe the mixture for a few minutes You will see a white threadlike substance rise from the mixture to rest above in the alcohol layer
This is the DNA that you have extracted from the cells of the kiwi fruit
8 You can get more DNA to precipitate from the solution using a DNA collecting tool (glass or paper clip hook or cut inoculation needle)
Gently lift the water solution up into the alcohol layer (this allows more DNA to get in contact with the alcohol and precipitate)
26
INFORMATION DNA precipitates as a white stringy
ldquosnottyrdquo film at the water - alcohol interface and eventually will rise into the alcohol layer from the mixture layer
Allow test tube to sit for several minutes The clearer the DNA is the fewer
impurities you have
If you have an acceptable amount of DNA it can be spooled by rotating your collecting tool and then transferred into a clean tube container
27
WOW ndash SPOOLING THE DNA If you are careful you may be able wind up the DNA
around a glass rod or a skewer Position the tip of the glass rod or skewer where you can see the threads of DNA Steadily twist the rod or skewer as if you were making candy floss Alternatively use a straw to pull it out by suction ndash be careful not to get in you mouth
Donrsquot go too quickly
You should be able to pull the strands of DNA out of the mixture
ASK STUDENTS TO TAKE MOBILE PHONE PICTURES OF THEIR OWN EXTRACTED DNA AND COMPARE APPEARANCE WITH OTHERS
Photos of steps can be inserted in flow diagram(ICT)
28
KIWIDNA
29
If you want to save your DNA you can transfer it to a small container filled with alcohol
Leave tube container uncapped until the ethanol has evaporated
DNA can be stored in the fridge (dry or water buffer can be added)
You can now investigate the property
30
WHAT IS THAT STRINGY STUFFDNA IS A LONG STRINGY MOLECULE THE SALT THAT YOU ADDED IN STEP ONE HELPS IT STICK TOGETHER SO WHAT YOU SEE ARE CLUMPS OF TANGLED DNA MOLECULES
DNA NORMALLY STAYS DISSOLVED IN WATER BUT WHEN SALTY DNA COMES IN CONTACT WITH ALCOHOL IT BECOMES UNDISSOLVED THIS IS CALLED PRECIPITATION THE PHYSICAL FORCE OF THE DNA CLUMPING TOGETHER AS IT PRECIPITATES PULLS MORE STRANDS ALONG WITH IT AS IT RISES INTO THE ALCOHOL
31
THE WHYrsquoS Blending separated the pea cells In order to extract DNA from a cell the associated
membranes and proteins must first be removed (break apart the cells) and then physically separated (loosen the tough cell wall) from the DNA
To see the DNA we have to break open these two sacks We do this with detergent and salt( Sodium can be
involved in several of the steps ) Sodium is an element Its chemical symbol is Na for
Natrium the Latin word for sodium It is a positive ion and often associates with negative ions as part of useful compounds
Salt help precipitate protein and carbohydrates away from the DNA
Salt helps strip away the proteins associated with DNA + and ndash charge
32
THE WHYrsquoS Salt and Detergents are used to break down
cell walls and nuclear membranes to release the DNA
They work by chemically poking holes in the cell membranes or walls
Once holes are poked in the membranes the membranes can be further disrupted mechanically as with a blender
After that it is easier to get the contents of the cell out including the DNA
LETS HAVE A LOOK AT THE CELL
33
CELL TO DNA
httplearngeneticsutaheducontentlabsextractionhowtoHOW TO EXTRACT DNA FROM ANYTHING LIVING
34
THE WHYrsquoS WHY DETERGENT
Each cell is surrounded by a sack (the cell membrane)
DNA is found inside the second sack (nucleus) within each cell
To see the DNA we have to break it open A cell membrane has 2 layers of lipid(fat)
molecules with protein going through them When the lysis buffer (detergent) comes
close to the cell it captures the lipids and the proteins - breaks open the cell destroying the fatty membrane - DNA now released into the solution
35
A CELLS MEMBRANES HAVE TWO LAYERS OF LIPID (FAT) MOLECULES WITH PROTEINS GOING THROUGH THEM
36
Why detergent How does detergent work
Think about why you use soap to wash dishes or your hands To remove grease and dirt right
Soap molecules and grease molecules are made of two parts
1(Blue) Heads which like water 2(Green) Tails which hate water
37
AFTER ADDING THE DETERGENT WHAT DO YOU HAVE IN YOUR SOUP
38
THE WHYrsquoS ENZYME (MEAT TENDERIZER)
The DNA in the nucleus of the cell is molded folded and protected by proteins The tenderizer cut the proteins away from the DNA
In this experiment meat tenderizer acts as an enzyme to cut proteins just like a pair of scissors
The meat tenderizer cuts the proteins away from the DNA
39
THE WHYrsquoS CUTTING PROTEIN AND DNA
The DNA in the nucleus of the cell is moulded folded and protected by proteins
40
THE WHYrsquoSALCOHOL Alcohol is less dense than water so it floats
on top Look for clumps of white stringy stuff where the water and alcohol layers meet
DNA is not soluble in alcohol - other cell parts are
By adding alcohol DNA precipitates out of the solution and collect at the interface of the alcohol and soap layer
The colder the alcohol the less soluble the DNA will be in it
41
COLD ALCOHOL
DNA dissolves in water but precipitates in alcohol
Cold alcohol is used to separate DNA out of water-based solutions
This allows the DNA to be purified for subsequent genetic testing
Adding alcohol to a solution containing DNA is a simple way to obtain the pure DNA required and colder temperatures slow down enzymes that can break down DNA giving better extraction results
42
WHY IS DNA EXTRACTION IMPORTANT DNA extraction is an important molecular
biology procedure
By definition extraction is taking DNA out of any type of cell for the purpose of analysis
See Handouts for more information on the use of DNA extracted (extension only)
43
SUMMARIZEDNA EXTRACTING FROM KIWI FRUIT
44
BREAK DOWN CELL WALLS
Cells walls have to be destroyed to reach the DNA within cells
Extraction procedures to obtain pure DNA have to get rid of all the molecular and chemical components of the tissue from which the DNA is being extracted
First steps involve lysing or destroying the cell walls This can be done with a variety of chemical agents that are caustic to cell membranes but do not harm the DNA Cells can also be sonicated homogenized or ground up to destroy membranes
45
REMOVING Removing Lipids
Once the cell walls have been destroyed a detergent is added to get rid of the fats and oils that make up the cell membranes Detergents cause the fats and oils to dissolve into the solution
Remove ProteinsProteins and enzymes can be digested by
adding a protease to the solution Proteases break down proteins into small peptides and amino acids
46
PRECIPITATING AND PURIFYING Precipitate DNA
Adding cold alcohol to a solution will cause DNA to precipitate and aggregate The DNA can be collected by centrifuging the sample and pouring off the liquid layer The DNA should exist as a small pellet in the bottom of the centrifuge tube
Purify DNAThe DNA can be washed by re-suspending it
in a cold alcohol solution and re-centrifuging it several times to obtain a very pure DNA sample Typical alcohols used to precipitate DNA include ethanol and Isopropanol This process leaves a very pure sample for stringent DNA testing
47
PART B OBSERVING EXTRACTED DNA UNDER THE MICROSCOPE
48
PART B A CLOSER LOOK
8 Use a dropping pipette to carefully remove some of the threadlike substance from the top of your preparation
9 Place one or two drops onto the middle of a
microscope slide
10 Add two drops of methylene blue Wait 3 or 4 minutes to allow the methylene blue to be absorbed by the DNA
11 Carefully place a cover slip on the slide Gently press
a folded piece of paper towelling over the top of the prepared slide to soak up any excess liquid
12 Observe the DNA under low power then high power
49
DISCUSSIONANSWER IN GROUP RELATED WORKED 1 Write a detailed description of the material
floating at the top of the test tube after the alcohol was added
2 Describe the DNA as it appears under the
microscope under high power 3 Prepare a diagram of your DNA specimen 4 What was the reason for using methylene
blue
50
HOMEWORKCONSTRUCT A FLOW CHART DRAWING MIND MAP OF THE PROCESS YOU USED EXTRACTING THE DNA FROM KIWIrsquoS
Next to each step explain how the method you used was important (Hint What substances make up the membranes of cells and cell organelles
How do detergents work
What is the active ingredient in meat tenderiser
51
TIPS STEPS FOR FLOW CHART In order to release the DNA from the
nuclei of the pea cells you first separated the cells from one another
Then the cell and nuclear membranes needed to be ruptured to release the cell contents and the contents of the nucleus
Once removed from the nuclear membrane the DNA had to be untangled into the visible threadlike structures you ended up with
52
HOMEWORK EXTENSION GROUP QUESTIONS Where can DNA be found in the cell Discuss the action of the soap (detergent) on the cell
What is the purpose of the soap in this activity What was the purpose of the Sodium Chloride Include a
discussion of polarity and charged particles Why was the cold ethanol added to the soap and salt
mixture Describe the appearance of your final product Draw a diagram of DNA containing 5 sets of nucleotide
bases labelling the hydrogen bonds between the bases References and Resources Adapted from Berry Full
of DNA by Diane Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
53
HTTPWWWGENOMEBCCAEDUCATIONTEACHERSCLASSROOM-ACTIVITIESDNA-EXTRACTION
54
EXTRACHROMOSOMES EXIST IN PAIRS Because our chromosomes exist in pairs (and consequently we
have 2 alleles of each gene) we are a diploid species This is why our somatic cells are represented as 2n Our gametes (sperm and ova) on the other hand are haploid and are represented as n Other species may have different ploidy for example
triploid (3n) seedless watermelons tetaploid (4n) salmonidae fish pentaploid (5n) Kenai birch hexaploid (6n) some types of wheat kiwi fruit octaploid (8n) acipenser (a genus
of sturgeon fish strawberies) decaploid (10n) some strawberries dodecaploid (12n) some types of amphibians eg Xenopus
ruwenzoriensis
wwwcyberusca
55
YOUR EXTRACTED DNA UNDER THE MICROSCOPE LOOKED
LIKE
Collaboration-Brainstorming on what to look at under the
- DNA extraction- DNA is too small for even a microscope to see and after the extraction it just appears like a blob True or false
Could you any DNA strand
wwwemployeescsbsjuedu
56
HANDOUT ON DNA EXTRACTIONBACKGROUND EVERY DAY LIFE
EXTRA READING MATERIAL(EXTENSION )
DISCOVERING DNA DNA ON THE INSIDE USE OF DNA EXTRACTION INN EVERYDAY LIFE WHY IS DNA TESTING GOOD MOLECULAR BIOLOGY PCR-based diagnostics of genomic DNA
57
REFERENCES httpwwwsquidoocomhow-to-get-dna-from-a-kiwi-fruit Why Is DNA Testing Good | eHowcom
httpwwwehowcomabout_6684410_dna-testing-good_htmlixzz1MkbcIJs5 Why Is DNA Extraction Important | eHowcom
httpwwwehowcomlist_5839095_dna-extraction-important_htmlixzz1MkZDrqyY
Why Is Sodium Used in DNA Extraction | eHowcom httpwwwehowcomabout_6504902_sodium-used-dna-extraction_htmlixzz1MkaGWg57
Why Is Cold Alcohol Used in DNA Tests | eHowcom httpwwwehowcomabout_6399349_cold-alcohol-used-dna-tests_htmlixzz1MkbEmEdu
httplearngeneticsutaheducontentlabsextractionhowto www kamibudaksainsblogspotcom
httpwwwchemwatch goldcom References and Resources Adapted from Berry Full of DNA by Diane
Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
58
REFERENCES
Uses of DNA Extraction | eHowcom httpwwwehowcomabout_5344428_uses-dna-extractionhtmlixzz1MkWQB2TZ
Science 10 A Contextual Approach Heineman Queensland Science Projects Regan Spence Maggie Spenceley
httpenwikipediaorgwikiMolecular_biology httpwwwclinchemorgcgicontent
extract44102201 http
employeescsbsjueduhjakubowskiclassesch331dnaoldnastructurehtml
httpwwwgenomebccaeducationteachersclassroom-activitiesdna-extraction
DO ONLY LIVING ORGANISMS CONTAIN DNA
What do you think
Discuss in pairs for 1 minute
Give reasons
5
EXTRACTING DNA FROM ANY LIVING THING
Just follow these 3 easy steps
DetergenteNzymes Alcohol
httplearngeneticsutaheducontentlabsextractionhowto
6
YO
U M
EA
N I
CA
N S
EE I
T
HO
W
DNA -
Deoxyribonucle
ic acid
bullAll living organisms contain DNA in their cells ndash FOUND in nucleus
bullDNA - The complex chemical compound found in chromosomes that
contains the genetic code
The nucleus is a membrane bound structure that contains the cells hereditary information and controls the cells growth and reproduction It is commonly the most prominent organelle in the cell
wwwkamibudaksainsblogspotcom
7
FIGURE HTTPEMPLOYEESCSBSJUEDUHJAKUBOWSKICLASSESCH331DNAOLDNASTRUCTUREHTML
PACKAGING OF DNA IN THE NUCLEUS
8
httpwwwyoutubecomwatchv=tmVP1cotozc
9
DNA extraction refers to the process that scientists use to break down a piece of evidence such as a piece of hair or a drop of blood in order to determine the individuals DNA which is unique to each person Understand the process of DNA extraction with information from a biology teacher in this free video on science
Expert Janice CrenettiContact WeAreHDTVcomBio Janice Creneti has a Bachelor of Science in secondary science education and a Bachelor of Art in biology from Boston UniversityFilmmaker Christopher Rokosz
Category Entertainment Tags science dna dna testing dna structure dna model dna replication dna molecule
10
REVIEWING OF INFORMATION ONLABORATORY SAFETY RULES AND RISK ASSESSMENTS
Do not enter the laboratory unless you are with a teacher Never touch equipment in the laboratory unless you are told to use it Donrsquot eat or drink in the laboratory Always walkmdashnever run Wear protective clothingmdash a laboratory coat or apron and when
appropriate safety glasses Never taste anything Tie up long hair Always point test tubes away from people Check with your teacher on how to dispose of waste liquids and solids
Broken glass should be cleaned up using gloves a brush and dustpan and placed in a special bin
If you spill something on your skin or clothes wash it immediately with lots of water Tell your teacher
Report all accidents and breakages to your teacher After heating equipment let it cool on a heatproof mat before picking
it up This will avoid burns Clean all equipment after use and put it back where you got it from
Clean and dry your work bench
11
RISK ASSESSMENT ON EXPERIMENT AND INVESTIGATION ndash KIWI FRUIT
Eyewear and protective clothingNo eat or drinkingCover the MSDSrsquo e on Isopropanol and Methylene blue ndash hard copy handouts (health and safety regulation)
Any broken glassware must be reported
Care must be taken when using and washing the blender ndash sharp blades
12
ISOPROPANOL ndash MSDS HAND OUTS CHEMWATCH ISSUE DATE 15052010 ISOPROPANOL - FLAMMABLE Dangerous Good 3 SAFETY-DATA(tm) Ratings (Provided here for your convenience)
----------------------------------------------------------------------------------------------------------- Health Rating 2 - Moderate Flammability Rating 3 - Severe (Flammable) Reactivity Rating 2 - Moderate Contact Rating 3 - Severe Lab Protective Equip GOGGLES amp SHIELD LAB COAT amp APRON VENT HOOD PROPER GLOVES CLASS B EXTINGUISHER Storage Colour Code Red (Flammable)
Potential Health Effects ----------------------------------
Inhalation Inhalation of vapours irritates the respiratory tract Exposure to high concentrations has a narcotic effect producing symptoms of dizziness drowsiness headache staggering unconsciousness and possibly death Ingestion Can cause drowsiness unconsciousness and death Gastrointestinal pain cramps nausea vomiting and diarrhoea may also result The single lethal dose for a human adult = about 250 mls (8 ounces) Skin Contact May cause irritation with redness and pain May be absorbed through the skin with possible systemic effects Eye Contact Vapours cause eye irritation Splashes cause severe irritation possible corneal burns and eye damage Chronic Exposure Chronic exposure may cause skin effects Aggravation of Pre-existing Conditions Persons with pre-existing skin disorders or impaired liver kidney or pulmonary function may be more susceptible to the effects of this agent
13
METHYLENE BLUE ndash MSDS HAND OUTS CHEMWATCH ISSUE DATE 15052010 METHYLENE BLUE SAFETY-DATA(tm) Ratings (Provided here for your convenience)
----------------------------------------------------------------------------------------------------------- Health Rating 2 - Moderate Flammability Rating 1 - Slight Reactivity Rating 1 - Slight Contact Rating 1 - Slight Lab Protective Equip GOGGLES LAB COAT VENT HOOD PROPER GLOVES Storage Colour Code Green (General Storage) -----------------------------------------------------------------------------------------------------------
Potential Health Effects ----------------------------------
This material is relatively nonhazardous in routine industrial situations
Inhalation No adverse health effects expected from inhalation May cause a short period of rapid or difficult breathing Ingestion A burning sensation of the mouth may be noted following ingestion of methylene blue May cause nausea vomiting diarrhea and gastritis Large doses may cause abdominal and chest pain headache profuse sweating mental confusion painful maturation and methemoglobinemia Skin Contact Not expected to be a health hazard from skin exposure Methylene blue may colour the skin a bluish colour May cause photosensitization Eye Contact No adverse effects expected May cause mechanical irritation Chronic Exposure No information found Aggravation of Pre-existing Conditions No information found
14
NEW WORDS TO WORD BANKADD THESE TO YOUR EXCEL ALPHA TABLE
Precipitate Spooling Denaturising Lysing Homogenized Aggregate Lipid Centrifuging PCR
WORDS MEANING
15
bullDO YOU THINK YOU HAVE VERY MUCH IN COMMON WITH A KIWI FRUIT
BELIEVE IT OR NOT A KIWIS GENETIC MATERIAL IS VERY SIMILAR TO YOUR OWN
SEE AND TOUCH THE GENETIC MATERIAL THAT YOULL EXTRACT FROM THE CELLS OF A KIWI FRUIT
Activity Overview
Extract DNA from kiwi fruit using simple household chemicals
16
HT
TP
LEA
RN
GEN
ETIC
SU
TA
HE
DU
CO
NTE
NTL
AB
SE
XTR
AC
TIO
NH
OW
TO
DNA EXTRACTION USING KIWI FRUIT
17
AIM
TO
EX
TR
AC
T D
NA
FR
OM
TH
E C
ELLS
OF K
IWI F
RU
IT
wwwexploratoriumedu
MATERIALS EQUIPMENT (per student group takes 30 minutes)
frac12 cup Kiwi fruit 200 mL water dishwashing detergent dropping pipette fine mesh kitchen strainer amp cheese cloth glass rod large beaker large test tube light microscope meat tenderiser methylene blue microscope lamp inoculation needle microscope slide and cover slip paper towelling small beaker of alcohol (Isopropanol) spatula test-tube rack mortar and pestle
18
PART A EXTRACTING THE DNA
METHOD
1 Peal and Place the kiwi and water in the beaker and mush it with a mortar
and pestle DNA SOURCE About 125 ml 12 cup Twice as much water as DNA source (250
ml 1 cup) Table salt large pinch 1g 14 teaspoon Stir until the mixture is of a thin soupy
consistency
TAKE NOTES ndash NO TEXTBOOKS ALLOWED WHEN PERFORMING EXPERIMENT ndashMUST RECALL PROCEDURES METHOD
19
WHAT MUST I DO NOW
STRAIN THE DNA MIXTURE
DISHWASHING DETERGENT
20
2 Pour the thin cell mixture through the kitchen strainer (cheesecloth) into another large beaker
3 Measure 12 ml of the soup into
a small beakerAdd 2 ml of liquid detergentSwirl to mix stir thoroughly using a
glass rodLet mixture sit in container with
hot tap water (60 - 65degC) for 5 - 10 minutes
21
WHAT MUST I DO NOWMEAT TENDERIZER
ENZYME POWDERALCOHOL SEPARATION
wwwforumsoverclockerscomau
22
4 Add a pinch of enzyme (meat tenderizer) to mixture
(Using pineapple juice or contact lens cleaning solution will do the same as tenderizer)
Gently stir with toothpick skewer for 5 minutes continue stirring not too vigorously Be careful If you stir too hard youll break up the DNA making it harder to see
5 Quarter-fill a large test tube with the mixture
23
SLOWLY pour the same amount ice - cold (70 - 95 isopropyl or ethyl alcohol) into test tube pour alcohol down the side of the test tube
Tilting the test tube will make this easier to do
It forms a layer on top of the cell mixture
DO NOT MIX THE TWO LAYERS TOGETHER
Amount of alcohol and mixture should be the same
24
SEPARATION USING ALCOHOL
25
7 Observe the mixture for a few minutes You will see a white threadlike substance rise from the mixture to rest above in the alcohol layer
This is the DNA that you have extracted from the cells of the kiwi fruit
8 You can get more DNA to precipitate from the solution using a DNA collecting tool (glass or paper clip hook or cut inoculation needle)
Gently lift the water solution up into the alcohol layer (this allows more DNA to get in contact with the alcohol and precipitate)
26
INFORMATION DNA precipitates as a white stringy
ldquosnottyrdquo film at the water - alcohol interface and eventually will rise into the alcohol layer from the mixture layer
Allow test tube to sit for several minutes The clearer the DNA is the fewer
impurities you have
If you have an acceptable amount of DNA it can be spooled by rotating your collecting tool and then transferred into a clean tube container
27
WOW ndash SPOOLING THE DNA If you are careful you may be able wind up the DNA
around a glass rod or a skewer Position the tip of the glass rod or skewer where you can see the threads of DNA Steadily twist the rod or skewer as if you were making candy floss Alternatively use a straw to pull it out by suction ndash be careful not to get in you mouth
Donrsquot go too quickly
You should be able to pull the strands of DNA out of the mixture
ASK STUDENTS TO TAKE MOBILE PHONE PICTURES OF THEIR OWN EXTRACTED DNA AND COMPARE APPEARANCE WITH OTHERS
Photos of steps can be inserted in flow diagram(ICT)
28
KIWIDNA
29
If you want to save your DNA you can transfer it to a small container filled with alcohol
Leave tube container uncapped until the ethanol has evaporated
DNA can be stored in the fridge (dry or water buffer can be added)
You can now investigate the property
30
WHAT IS THAT STRINGY STUFFDNA IS A LONG STRINGY MOLECULE THE SALT THAT YOU ADDED IN STEP ONE HELPS IT STICK TOGETHER SO WHAT YOU SEE ARE CLUMPS OF TANGLED DNA MOLECULES
DNA NORMALLY STAYS DISSOLVED IN WATER BUT WHEN SALTY DNA COMES IN CONTACT WITH ALCOHOL IT BECOMES UNDISSOLVED THIS IS CALLED PRECIPITATION THE PHYSICAL FORCE OF THE DNA CLUMPING TOGETHER AS IT PRECIPITATES PULLS MORE STRANDS ALONG WITH IT AS IT RISES INTO THE ALCOHOL
31
THE WHYrsquoS Blending separated the pea cells In order to extract DNA from a cell the associated
membranes and proteins must first be removed (break apart the cells) and then physically separated (loosen the tough cell wall) from the DNA
To see the DNA we have to break open these two sacks We do this with detergent and salt( Sodium can be
involved in several of the steps ) Sodium is an element Its chemical symbol is Na for
Natrium the Latin word for sodium It is a positive ion and often associates with negative ions as part of useful compounds
Salt help precipitate protein and carbohydrates away from the DNA
Salt helps strip away the proteins associated with DNA + and ndash charge
32
THE WHYrsquoS Salt and Detergents are used to break down
cell walls and nuclear membranes to release the DNA
They work by chemically poking holes in the cell membranes or walls
Once holes are poked in the membranes the membranes can be further disrupted mechanically as with a blender
After that it is easier to get the contents of the cell out including the DNA
LETS HAVE A LOOK AT THE CELL
33
CELL TO DNA
httplearngeneticsutaheducontentlabsextractionhowtoHOW TO EXTRACT DNA FROM ANYTHING LIVING
34
THE WHYrsquoS WHY DETERGENT
Each cell is surrounded by a sack (the cell membrane)
DNA is found inside the second sack (nucleus) within each cell
To see the DNA we have to break it open A cell membrane has 2 layers of lipid(fat)
molecules with protein going through them When the lysis buffer (detergent) comes
close to the cell it captures the lipids and the proteins - breaks open the cell destroying the fatty membrane - DNA now released into the solution
35
A CELLS MEMBRANES HAVE TWO LAYERS OF LIPID (FAT) MOLECULES WITH PROTEINS GOING THROUGH THEM
36
Why detergent How does detergent work
Think about why you use soap to wash dishes or your hands To remove grease and dirt right
Soap molecules and grease molecules are made of two parts
1(Blue) Heads which like water 2(Green) Tails which hate water
37
AFTER ADDING THE DETERGENT WHAT DO YOU HAVE IN YOUR SOUP
38
THE WHYrsquoS ENZYME (MEAT TENDERIZER)
The DNA in the nucleus of the cell is molded folded and protected by proteins The tenderizer cut the proteins away from the DNA
In this experiment meat tenderizer acts as an enzyme to cut proteins just like a pair of scissors
The meat tenderizer cuts the proteins away from the DNA
39
THE WHYrsquoS CUTTING PROTEIN AND DNA
The DNA in the nucleus of the cell is moulded folded and protected by proteins
40
THE WHYrsquoSALCOHOL Alcohol is less dense than water so it floats
on top Look for clumps of white stringy stuff where the water and alcohol layers meet
DNA is not soluble in alcohol - other cell parts are
By adding alcohol DNA precipitates out of the solution and collect at the interface of the alcohol and soap layer
The colder the alcohol the less soluble the DNA will be in it
41
COLD ALCOHOL
DNA dissolves in water but precipitates in alcohol
Cold alcohol is used to separate DNA out of water-based solutions
This allows the DNA to be purified for subsequent genetic testing
Adding alcohol to a solution containing DNA is a simple way to obtain the pure DNA required and colder temperatures slow down enzymes that can break down DNA giving better extraction results
42
WHY IS DNA EXTRACTION IMPORTANT DNA extraction is an important molecular
biology procedure
By definition extraction is taking DNA out of any type of cell for the purpose of analysis
See Handouts for more information on the use of DNA extracted (extension only)
43
SUMMARIZEDNA EXTRACTING FROM KIWI FRUIT
44
BREAK DOWN CELL WALLS
Cells walls have to be destroyed to reach the DNA within cells
Extraction procedures to obtain pure DNA have to get rid of all the molecular and chemical components of the tissue from which the DNA is being extracted
First steps involve lysing or destroying the cell walls This can be done with a variety of chemical agents that are caustic to cell membranes but do not harm the DNA Cells can also be sonicated homogenized or ground up to destroy membranes
45
REMOVING Removing Lipids
Once the cell walls have been destroyed a detergent is added to get rid of the fats and oils that make up the cell membranes Detergents cause the fats and oils to dissolve into the solution
Remove ProteinsProteins and enzymes can be digested by
adding a protease to the solution Proteases break down proteins into small peptides and amino acids
46
PRECIPITATING AND PURIFYING Precipitate DNA
Adding cold alcohol to a solution will cause DNA to precipitate and aggregate The DNA can be collected by centrifuging the sample and pouring off the liquid layer The DNA should exist as a small pellet in the bottom of the centrifuge tube
Purify DNAThe DNA can be washed by re-suspending it
in a cold alcohol solution and re-centrifuging it several times to obtain a very pure DNA sample Typical alcohols used to precipitate DNA include ethanol and Isopropanol This process leaves a very pure sample for stringent DNA testing
47
PART B OBSERVING EXTRACTED DNA UNDER THE MICROSCOPE
48
PART B A CLOSER LOOK
8 Use a dropping pipette to carefully remove some of the threadlike substance from the top of your preparation
9 Place one or two drops onto the middle of a
microscope slide
10 Add two drops of methylene blue Wait 3 or 4 minutes to allow the methylene blue to be absorbed by the DNA
11 Carefully place a cover slip on the slide Gently press
a folded piece of paper towelling over the top of the prepared slide to soak up any excess liquid
12 Observe the DNA under low power then high power
49
DISCUSSIONANSWER IN GROUP RELATED WORKED 1 Write a detailed description of the material
floating at the top of the test tube after the alcohol was added
2 Describe the DNA as it appears under the
microscope under high power 3 Prepare a diagram of your DNA specimen 4 What was the reason for using methylene
blue
50
HOMEWORKCONSTRUCT A FLOW CHART DRAWING MIND MAP OF THE PROCESS YOU USED EXTRACTING THE DNA FROM KIWIrsquoS
Next to each step explain how the method you used was important (Hint What substances make up the membranes of cells and cell organelles
How do detergents work
What is the active ingredient in meat tenderiser
51
TIPS STEPS FOR FLOW CHART In order to release the DNA from the
nuclei of the pea cells you first separated the cells from one another
Then the cell and nuclear membranes needed to be ruptured to release the cell contents and the contents of the nucleus
Once removed from the nuclear membrane the DNA had to be untangled into the visible threadlike structures you ended up with
52
HOMEWORK EXTENSION GROUP QUESTIONS Where can DNA be found in the cell Discuss the action of the soap (detergent) on the cell
What is the purpose of the soap in this activity What was the purpose of the Sodium Chloride Include a
discussion of polarity and charged particles Why was the cold ethanol added to the soap and salt
mixture Describe the appearance of your final product Draw a diagram of DNA containing 5 sets of nucleotide
bases labelling the hydrogen bonds between the bases References and Resources Adapted from Berry Full
of DNA by Diane Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
53
HTTPWWWGENOMEBCCAEDUCATIONTEACHERSCLASSROOM-ACTIVITIESDNA-EXTRACTION
54
EXTRACHROMOSOMES EXIST IN PAIRS Because our chromosomes exist in pairs (and consequently we
have 2 alleles of each gene) we are a diploid species This is why our somatic cells are represented as 2n Our gametes (sperm and ova) on the other hand are haploid and are represented as n Other species may have different ploidy for example
triploid (3n) seedless watermelons tetaploid (4n) salmonidae fish pentaploid (5n) Kenai birch hexaploid (6n) some types of wheat kiwi fruit octaploid (8n) acipenser (a genus
of sturgeon fish strawberies) decaploid (10n) some strawberries dodecaploid (12n) some types of amphibians eg Xenopus
ruwenzoriensis
wwwcyberusca
55
YOUR EXTRACTED DNA UNDER THE MICROSCOPE LOOKED
LIKE
Collaboration-Brainstorming on what to look at under the
- DNA extraction- DNA is too small for even a microscope to see and after the extraction it just appears like a blob True or false
Could you any DNA strand
wwwemployeescsbsjuedu
56
HANDOUT ON DNA EXTRACTIONBACKGROUND EVERY DAY LIFE
EXTRA READING MATERIAL(EXTENSION )
DISCOVERING DNA DNA ON THE INSIDE USE OF DNA EXTRACTION INN EVERYDAY LIFE WHY IS DNA TESTING GOOD MOLECULAR BIOLOGY PCR-based diagnostics of genomic DNA
57
REFERENCES httpwwwsquidoocomhow-to-get-dna-from-a-kiwi-fruit Why Is DNA Testing Good | eHowcom
httpwwwehowcomabout_6684410_dna-testing-good_htmlixzz1MkbcIJs5 Why Is DNA Extraction Important | eHowcom
httpwwwehowcomlist_5839095_dna-extraction-important_htmlixzz1MkZDrqyY
Why Is Sodium Used in DNA Extraction | eHowcom httpwwwehowcomabout_6504902_sodium-used-dna-extraction_htmlixzz1MkaGWg57
Why Is Cold Alcohol Used in DNA Tests | eHowcom httpwwwehowcomabout_6399349_cold-alcohol-used-dna-tests_htmlixzz1MkbEmEdu
httplearngeneticsutaheducontentlabsextractionhowto www kamibudaksainsblogspotcom
httpwwwchemwatch goldcom References and Resources Adapted from Berry Full of DNA by Diane
Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
58
REFERENCES
Uses of DNA Extraction | eHowcom httpwwwehowcomabout_5344428_uses-dna-extractionhtmlixzz1MkWQB2TZ
Science 10 A Contextual Approach Heineman Queensland Science Projects Regan Spence Maggie Spenceley
httpenwikipediaorgwikiMolecular_biology httpwwwclinchemorgcgicontent
extract44102201 http
employeescsbsjueduhjakubowskiclassesch331dnaoldnastructurehtml
httpwwwgenomebccaeducationteachersclassroom-activitiesdna-extraction
5
EXTRACTING DNA FROM ANY LIVING THING
Just follow these 3 easy steps
DetergenteNzymes Alcohol
httplearngeneticsutaheducontentlabsextractionhowto
6
YO
U M
EA
N I
CA
N S
EE I
T
HO
W
DNA -
Deoxyribonucle
ic acid
bullAll living organisms contain DNA in their cells ndash FOUND in nucleus
bullDNA - The complex chemical compound found in chromosomes that
contains the genetic code
The nucleus is a membrane bound structure that contains the cells hereditary information and controls the cells growth and reproduction It is commonly the most prominent organelle in the cell
wwwkamibudaksainsblogspotcom
7
FIGURE HTTPEMPLOYEESCSBSJUEDUHJAKUBOWSKICLASSESCH331DNAOLDNASTRUCTUREHTML
PACKAGING OF DNA IN THE NUCLEUS
8
httpwwwyoutubecomwatchv=tmVP1cotozc
9
DNA extraction refers to the process that scientists use to break down a piece of evidence such as a piece of hair or a drop of blood in order to determine the individuals DNA which is unique to each person Understand the process of DNA extraction with information from a biology teacher in this free video on science
Expert Janice CrenettiContact WeAreHDTVcomBio Janice Creneti has a Bachelor of Science in secondary science education and a Bachelor of Art in biology from Boston UniversityFilmmaker Christopher Rokosz
Category Entertainment Tags science dna dna testing dna structure dna model dna replication dna molecule
10
REVIEWING OF INFORMATION ONLABORATORY SAFETY RULES AND RISK ASSESSMENTS
Do not enter the laboratory unless you are with a teacher Never touch equipment in the laboratory unless you are told to use it Donrsquot eat or drink in the laboratory Always walkmdashnever run Wear protective clothingmdash a laboratory coat or apron and when
appropriate safety glasses Never taste anything Tie up long hair Always point test tubes away from people Check with your teacher on how to dispose of waste liquids and solids
Broken glass should be cleaned up using gloves a brush and dustpan and placed in a special bin
If you spill something on your skin or clothes wash it immediately with lots of water Tell your teacher
Report all accidents and breakages to your teacher After heating equipment let it cool on a heatproof mat before picking
it up This will avoid burns Clean all equipment after use and put it back where you got it from
Clean and dry your work bench
11
RISK ASSESSMENT ON EXPERIMENT AND INVESTIGATION ndash KIWI FRUIT
Eyewear and protective clothingNo eat or drinkingCover the MSDSrsquo e on Isopropanol and Methylene blue ndash hard copy handouts (health and safety regulation)
Any broken glassware must be reported
Care must be taken when using and washing the blender ndash sharp blades
12
ISOPROPANOL ndash MSDS HAND OUTS CHEMWATCH ISSUE DATE 15052010 ISOPROPANOL - FLAMMABLE Dangerous Good 3 SAFETY-DATA(tm) Ratings (Provided here for your convenience)
----------------------------------------------------------------------------------------------------------- Health Rating 2 - Moderate Flammability Rating 3 - Severe (Flammable) Reactivity Rating 2 - Moderate Contact Rating 3 - Severe Lab Protective Equip GOGGLES amp SHIELD LAB COAT amp APRON VENT HOOD PROPER GLOVES CLASS B EXTINGUISHER Storage Colour Code Red (Flammable)
Potential Health Effects ----------------------------------
Inhalation Inhalation of vapours irritates the respiratory tract Exposure to high concentrations has a narcotic effect producing symptoms of dizziness drowsiness headache staggering unconsciousness and possibly death Ingestion Can cause drowsiness unconsciousness and death Gastrointestinal pain cramps nausea vomiting and diarrhoea may also result The single lethal dose for a human adult = about 250 mls (8 ounces) Skin Contact May cause irritation with redness and pain May be absorbed through the skin with possible systemic effects Eye Contact Vapours cause eye irritation Splashes cause severe irritation possible corneal burns and eye damage Chronic Exposure Chronic exposure may cause skin effects Aggravation of Pre-existing Conditions Persons with pre-existing skin disorders or impaired liver kidney or pulmonary function may be more susceptible to the effects of this agent
13
METHYLENE BLUE ndash MSDS HAND OUTS CHEMWATCH ISSUE DATE 15052010 METHYLENE BLUE SAFETY-DATA(tm) Ratings (Provided here for your convenience)
----------------------------------------------------------------------------------------------------------- Health Rating 2 - Moderate Flammability Rating 1 - Slight Reactivity Rating 1 - Slight Contact Rating 1 - Slight Lab Protective Equip GOGGLES LAB COAT VENT HOOD PROPER GLOVES Storage Colour Code Green (General Storage) -----------------------------------------------------------------------------------------------------------
Potential Health Effects ----------------------------------
This material is relatively nonhazardous in routine industrial situations
Inhalation No adverse health effects expected from inhalation May cause a short period of rapid or difficult breathing Ingestion A burning sensation of the mouth may be noted following ingestion of methylene blue May cause nausea vomiting diarrhea and gastritis Large doses may cause abdominal and chest pain headache profuse sweating mental confusion painful maturation and methemoglobinemia Skin Contact Not expected to be a health hazard from skin exposure Methylene blue may colour the skin a bluish colour May cause photosensitization Eye Contact No adverse effects expected May cause mechanical irritation Chronic Exposure No information found Aggravation of Pre-existing Conditions No information found
14
NEW WORDS TO WORD BANKADD THESE TO YOUR EXCEL ALPHA TABLE
Precipitate Spooling Denaturising Lysing Homogenized Aggregate Lipid Centrifuging PCR
WORDS MEANING
15
bullDO YOU THINK YOU HAVE VERY MUCH IN COMMON WITH A KIWI FRUIT
BELIEVE IT OR NOT A KIWIS GENETIC MATERIAL IS VERY SIMILAR TO YOUR OWN
SEE AND TOUCH THE GENETIC MATERIAL THAT YOULL EXTRACT FROM THE CELLS OF A KIWI FRUIT
Activity Overview
Extract DNA from kiwi fruit using simple household chemicals
16
HT
TP
LEA
RN
GEN
ETIC
SU
TA
HE
DU
CO
NTE
NTL
AB
SE
XTR
AC
TIO
NH
OW
TO
DNA EXTRACTION USING KIWI FRUIT
17
AIM
TO
EX
TR
AC
T D
NA
FR
OM
TH
E C
ELLS
OF K
IWI F
RU
IT
wwwexploratoriumedu
MATERIALS EQUIPMENT (per student group takes 30 minutes)
frac12 cup Kiwi fruit 200 mL water dishwashing detergent dropping pipette fine mesh kitchen strainer amp cheese cloth glass rod large beaker large test tube light microscope meat tenderiser methylene blue microscope lamp inoculation needle microscope slide and cover slip paper towelling small beaker of alcohol (Isopropanol) spatula test-tube rack mortar and pestle
18
PART A EXTRACTING THE DNA
METHOD
1 Peal and Place the kiwi and water in the beaker and mush it with a mortar
and pestle DNA SOURCE About 125 ml 12 cup Twice as much water as DNA source (250
ml 1 cup) Table salt large pinch 1g 14 teaspoon Stir until the mixture is of a thin soupy
consistency
TAKE NOTES ndash NO TEXTBOOKS ALLOWED WHEN PERFORMING EXPERIMENT ndashMUST RECALL PROCEDURES METHOD
19
WHAT MUST I DO NOW
STRAIN THE DNA MIXTURE
DISHWASHING DETERGENT
20
2 Pour the thin cell mixture through the kitchen strainer (cheesecloth) into another large beaker
3 Measure 12 ml of the soup into
a small beakerAdd 2 ml of liquid detergentSwirl to mix stir thoroughly using a
glass rodLet mixture sit in container with
hot tap water (60 - 65degC) for 5 - 10 minutes
21
WHAT MUST I DO NOWMEAT TENDERIZER
ENZYME POWDERALCOHOL SEPARATION
wwwforumsoverclockerscomau
22
4 Add a pinch of enzyme (meat tenderizer) to mixture
(Using pineapple juice or contact lens cleaning solution will do the same as tenderizer)
Gently stir with toothpick skewer for 5 minutes continue stirring not too vigorously Be careful If you stir too hard youll break up the DNA making it harder to see
5 Quarter-fill a large test tube with the mixture
23
SLOWLY pour the same amount ice - cold (70 - 95 isopropyl or ethyl alcohol) into test tube pour alcohol down the side of the test tube
Tilting the test tube will make this easier to do
It forms a layer on top of the cell mixture
DO NOT MIX THE TWO LAYERS TOGETHER
Amount of alcohol and mixture should be the same
24
SEPARATION USING ALCOHOL
25
7 Observe the mixture for a few minutes You will see a white threadlike substance rise from the mixture to rest above in the alcohol layer
This is the DNA that you have extracted from the cells of the kiwi fruit
8 You can get more DNA to precipitate from the solution using a DNA collecting tool (glass or paper clip hook or cut inoculation needle)
Gently lift the water solution up into the alcohol layer (this allows more DNA to get in contact with the alcohol and precipitate)
26
INFORMATION DNA precipitates as a white stringy
ldquosnottyrdquo film at the water - alcohol interface and eventually will rise into the alcohol layer from the mixture layer
Allow test tube to sit for several minutes The clearer the DNA is the fewer
impurities you have
If you have an acceptable amount of DNA it can be spooled by rotating your collecting tool and then transferred into a clean tube container
27
WOW ndash SPOOLING THE DNA If you are careful you may be able wind up the DNA
around a glass rod or a skewer Position the tip of the glass rod or skewer where you can see the threads of DNA Steadily twist the rod or skewer as if you were making candy floss Alternatively use a straw to pull it out by suction ndash be careful not to get in you mouth
Donrsquot go too quickly
You should be able to pull the strands of DNA out of the mixture
ASK STUDENTS TO TAKE MOBILE PHONE PICTURES OF THEIR OWN EXTRACTED DNA AND COMPARE APPEARANCE WITH OTHERS
Photos of steps can be inserted in flow diagram(ICT)
28
KIWIDNA
29
If you want to save your DNA you can transfer it to a small container filled with alcohol
Leave tube container uncapped until the ethanol has evaporated
DNA can be stored in the fridge (dry or water buffer can be added)
You can now investigate the property
30
WHAT IS THAT STRINGY STUFFDNA IS A LONG STRINGY MOLECULE THE SALT THAT YOU ADDED IN STEP ONE HELPS IT STICK TOGETHER SO WHAT YOU SEE ARE CLUMPS OF TANGLED DNA MOLECULES
DNA NORMALLY STAYS DISSOLVED IN WATER BUT WHEN SALTY DNA COMES IN CONTACT WITH ALCOHOL IT BECOMES UNDISSOLVED THIS IS CALLED PRECIPITATION THE PHYSICAL FORCE OF THE DNA CLUMPING TOGETHER AS IT PRECIPITATES PULLS MORE STRANDS ALONG WITH IT AS IT RISES INTO THE ALCOHOL
31
THE WHYrsquoS Blending separated the pea cells In order to extract DNA from a cell the associated
membranes and proteins must first be removed (break apart the cells) and then physically separated (loosen the tough cell wall) from the DNA
To see the DNA we have to break open these two sacks We do this with detergent and salt( Sodium can be
involved in several of the steps ) Sodium is an element Its chemical symbol is Na for
Natrium the Latin word for sodium It is a positive ion and often associates with negative ions as part of useful compounds
Salt help precipitate protein and carbohydrates away from the DNA
Salt helps strip away the proteins associated with DNA + and ndash charge
32
THE WHYrsquoS Salt and Detergents are used to break down
cell walls and nuclear membranes to release the DNA
They work by chemically poking holes in the cell membranes or walls
Once holes are poked in the membranes the membranes can be further disrupted mechanically as with a blender
After that it is easier to get the contents of the cell out including the DNA
LETS HAVE A LOOK AT THE CELL
33
CELL TO DNA
httplearngeneticsutaheducontentlabsextractionhowtoHOW TO EXTRACT DNA FROM ANYTHING LIVING
34
THE WHYrsquoS WHY DETERGENT
Each cell is surrounded by a sack (the cell membrane)
DNA is found inside the second sack (nucleus) within each cell
To see the DNA we have to break it open A cell membrane has 2 layers of lipid(fat)
molecules with protein going through them When the lysis buffer (detergent) comes
close to the cell it captures the lipids and the proteins - breaks open the cell destroying the fatty membrane - DNA now released into the solution
35
A CELLS MEMBRANES HAVE TWO LAYERS OF LIPID (FAT) MOLECULES WITH PROTEINS GOING THROUGH THEM
36
Why detergent How does detergent work
Think about why you use soap to wash dishes or your hands To remove grease and dirt right
Soap molecules and grease molecules are made of two parts
1(Blue) Heads which like water 2(Green) Tails which hate water
37
AFTER ADDING THE DETERGENT WHAT DO YOU HAVE IN YOUR SOUP
38
THE WHYrsquoS ENZYME (MEAT TENDERIZER)
The DNA in the nucleus of the cell is molded folded and protected by proteins The tenderizer cut the proteins away from the DNA
In this experiment meat tenderizer acts as an enzyme to cut proteins just like a pair of scissors
The meat tenderizer cuts the proteins away from the DNA
39
THE WHYrsquoS CUTTING PROTEIN AND DNA
The DNA in the nucleus of the cell is moulded folded and protected by proteins
40
THE WHYrsquoSALCOHOL Alcohol is less dense than water so it floats
on top Look for clumps of white stringy stuff where the water and alcohol layers meet
DNA is not soluble in alcohol - other cell parts are
By adding alcohol DNA precipitates out of the solution and collect at the interface of the alcohol and soap layer
The colder the alcohol the less soluble the DNA will be in it
41
COLD ALCOHOL
DNA dissolves in water but precipitates in alcohol
Cold alcohol is used to separate DNA out of water-based solutions
This allows the DNA to be purified for subsequent genetic testing
Adding alcohol to a solution containing DNA is a simple way to obtain the pure DNA required and colder temperatures slow down enzymes that can break down DNA giving better extraction results
42
WHY IS DNA EXTRACTION IMPORTANT DNA extraction is an important molecular
biology procedure
By definition extraction is taking DNA out of any type of cell for the purpose of analysis
See Handouts for more information on the use of DNA extracted (extension only)
43
SUMMARIZEDNA EXTRACTING FROM KIWI FRUIT
44
BREAK DOWN CELL WALLS
Cells walls have to be destroyed to reach the DNA within cells
Extraction procedures to obtain pure DNA have to get rid of all the molecular and chemical components of the tissue from which the DNA is being extracted
First steps involve lysing or destroying the cell walls This can be done with a variety of chemical agents that are caustic to cell membranes but do not harm the DNA Cells can also be sonicated homogenized or ground up to destroy membranes
45
REMOVING Removing Lipids
Once the cell walls have been destroyed a detergent is added to get rid of the fats and oils that make up the cell membranes Detergents cause the fats and oils to dissolve into the solution
Remove ProteinsProteins and enzymes can be digested by
adding a protease to the solution Proteases break down proteins into small peptides and amino acids
46
PRECIPITATING AND PURIFYING Precipitate DNA
Adding cold alcohol to a solution will cause DNA to precipitate and aggregate The DNA can be collected by centrifuging the sample and pouring off the liquid layer The DNA should exist as a small pellet in the bottom of the centrifuge tube
Purify DNAThe DNA can be washed by re-suspending it
in a cold alcohol solution and re-centrifuging it several times to obtain a very pure DNA sample Typical alcohols used to precipitate DNA include ethanol and Isopropanol This process leaves a very pure sample for stringent DNA testing
47
PART B OBSERVING EXTRACTED DNA UNDER THE MICROSCOPE
48
PART B A CLOSER LOOK
8 Use a dropping pipette to carefully remove some of the threadlike substance from the top of your preparation
9 Place one or two drops onto the middle of a
microscope slide
10 Add two drops of methylene blue Wait 3 or 4 minutes to allow the methylene blue to be absorbed by the DNA
11 Carefully place a cover slip on the slide Gently press
a folded piece of paper towelling over the top of the prepared slide to soak up any excess liquid
12 Observe the DNA under low power then high power
49
DISCUSSIONANSWER IN GROUP RELATED WORKED 1 Write a detailed description of the material
floating at the top of the test tube after the alcohol was added
2 Describe the DNA as it appears under the
microscope under high power 3 Prepare a diagram of your DNA specimen 4 What was the reason for using methylene
blue
50
HOMEWORKCONSTRUCT A FLOW CHART DRAWING MIND MAP OF THE PROCESS YOU USED EXTRACTING THE DNA FROM KIWIrsquoS
Next to each step explain how the method you used was important (Hint What substances make up the membranes of cells and cell organelles
How do detergents work
What is the active ingredient in meat tenderiser
51
TIPS STEPS FOR FLOW CHART In order to release the DNA from the
nuclei of the pea cells you first separated the cells from one another
Then the cell and nuclear membranes needed to be ruptured to release the cell contents and the contents of the nucleus
Once removed from the nuclear membrane the DNA had to be untangled into the visible threadlike structures you ended up with
52
HOMEWORK EXTENSION GROUP QUESTIONS Where can DNA be found in the cell Discuss the action of the soap (detergent) on the cell
What is the purpose of the soap in this activity What was the purpose of the Sodium Chloride Include a
discussion of polarity and charged particles Why was the cold ethanol added to the soap and salt
mixture Describe the appearance of your final product Draw a diagram of DNA containing 5 sets of nucleotide
bases labelling the hydrogen bonds between the bases References and Resources Adapted from Berry Full
of DNA by Diane Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
53
HTTPWWWGENOMEBCCAEDUCATIONTEACHERSCLASSROOM-ACTIVITIESDNA-EXTRACTION
54
EXTRACHROMOSOMES EXIST IN PAIRS Because our chromosomes exist in pairs (and consequently we
have 2 alleles of each gene) we are a diploid species This is why our somatic cells are represented as 2n Our gametes (sperm and ova) on the other hand are haploid and are represented as n Other species may have different ploidy for example
triploid (3n) seedless watermelons tetaploid (4n) salmonidae fish pentaploid (5n) Kenai birch hexaploid (6n) some types of wheat kiwi fruit octaploid (8n) acipenser (a genus
of sturgeon fish strawberies) decaploid (10n) some strawberries dodecaploid (12n) some types of amphibians eg Xenopus
ruwenzoriensis
wwwcyberusca
55
YOUR EXTRACTED DNA UNDER THE MICROSCOPE LOOKED
LIKE
Collaboration-Brainstorming on what to look at under the
- DNA extraction- DNA is too small for even a microscope to see and after the extraction it just appears like a blob True or false
Could you any DNA strand
wwwemployeescsbsjuedu
56
HANDOUT ON DNA EXTRACTIONBACKGROUND EVERY DAY LIFE
EXTRA READING MATERIAL(EXTENSION )
DISCOVERING DNA DNA ON THE INSIDE USE OF DNA EXTRACTION INN EVERYDAY LIFE WHY IS DNA TESTING GOOD MOLECULAR BIOLOGY PCR-based diagnostics of genomic DNA
57
REFERENCES httpwwwsquidoocomhow-to-get-dna-from-a-kiwi-fruit Why Is DNA Testing Good | eHowcom
httpwwwehowcomabout_6684410_dna-testing-good_htmlixzz1MkbcIJs5 Why Is DNA Extraction Important | eHowcom
httpwwwehowcomlist_5839095_dna-extraction-important_htmlixzz1MkZDrqyY
Why Is Sodium Used in DNA Extraction | eHowcom httpwwwehowcomabout_6504902_sodium-used-dna-extraction_htmlixzz1MkaGWg57
Why Is Cold Alcohol Used in DNA Tests | eHowcom httpwwwehowcomabout_6399349_cold-alcohol-used-dna-tests_htmlixzz1MkbEmEdu
httplearngeneticsutaheducontentlabsextractionhowto www kamibudaksainsblogspotcom
httpwwwchemwatch goldcom References and Resources Adapted from Berry Full of DNA by Diane
Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
58
REFERENCES
Uses of DNA Extraction | eHowcom httpwwwehowcomabout_5344428_uses-dna-extractionhtmlixzz1MkWQB2TZ
Science 10 A Contextual Approach Heineman Queensland Science Projects Regan Spence Maggie Spenceley
httpenwikipediaorgwikiMolecular_biology httpwwwclinchemorgcgicontent
extract44102201 http
employeescsbsjueduhjakubowskiclassesch331dnaoldnastructurehtml
httpwwwgenomebccaeducationteachersclassroom-activitiesdna-extraction
6
YO
U M
EA
N I
CA
N S
EE I
T
HO
W
DNA -
Deoxyribonucle
ic acid
bullAll living organisms contain DNA in their cells ndash FOUND in nucleus
bullDNA - The complex chemical compound found in chromosomes that
contains the genetic code
The nucleus is a membrane bound structure that contains the cells hereditary information and controls the cells growth and reproduction It is commonly the most prominent organelle in the cell
wwwkamibudaksainsblogspotcom
7
FIGURE HTTPEMPLOYEESCSBSJUEDUHJAKUBOWSKICLASSESCH331DNAOLDNASTRUCTUREHTML
PACKAGING OF DNA IN THE NUCLEUS
8
httpwwwyoutubecomwatchv=tmVP1cotozc
9
DNA extraction refers to the process that scientists use to break down a piece of evidence such as a piece of hair or a drop of blood in order to determine the individuals DNA which is unique to each person Understand the process of DNA extraction with information from a biology teacher in this free video on science
Expert Janice CrenettiContact WeAreHDTVcomBio Janice Creneti has a Bachelor of Science in secondary science education and a Bachelor of Art in biology from Boston UniversityFilmmaker Christopher Rokosz
Category Entertainment Tags science dna dna testing dna structure dna model dna replication dna molecule
10
REVIEWING OF INFORMATION ONLABORATORY SAFETY RULES AND RISK ASSESSMENTS
Do not enter the laboratory unless you are with a teacher Never touch equipment in the laboratory unless you are told to use it Donrsquot eat or drink in the laboratory Always walkmdashnever run Wear protective clothingmdash a laboratory coat or apron and when
appropriate safety glasses Never taste anything Tie up long hair Always point test tubes away from people Check with your teacher on how to dispose of waste liquids and solids
Broken glass should be cleaned up using gloves a brush and dustpan and placed in a special bin
If you spill something on your skin or clothes wash it immediately with lots of water Tell your teacher
Report all accidents and breakages to your teacher After heating equipment let it cool on a heatproof mat before picking
it up This will avoid burns Clean all equipment after use and put it back where you got it from
Clean and dry your work bench
11
RISK ASSESSMENT ON EXPERIMENT AND INVESTIGATION ndash KIWI FRUIT
Eyewear and protective clothingNo eat or drinkingCover the MSDSrsquo e on Isopropanol and Methylene blue ndash hard copy handouts (health and safety regulation)
Any broken glassware must be reported
Care must be taken when using and washing the blender ndash sharp blades
12
ISOPROPANOL ndash MSDS HAND OUTS CHEMWATCH ISSUE DATE 15052010 ISOPROPANOL - FLAMMABLE Dangerous Good 3 SAFETY-DATA(tm) Ratings (Provided here for your convenience)
----------------------------------------------------------------------------------------------------------- Health Rating 2 - Moderate Flammability Rating 3 - Severe (Flammable) Reactivity Rating 2 - Moderate Contact Rating 3 - Severe Lab Protective Equip GOGGLES amp SHIELD LAB COAT amp APRON VENT HOOD PROPER GLOVES CLASS B EXTINGUISHER Storage Colour Code Red (Flammable)
Potential Health Effects ----------------------------------
Inhalation Inhalation of vapours irritates the respiratory tract Exposure to high concentrations has a narcotic effect producing symptoms of dizziness drowsiness headache staggering unconsciousness and possibly death Ingestion Can cause drowsiness unconsciousness and death Gastrointestinal pain cramps nausea vomiting and diarrhoea may also result The single lethal dose for a human adult = about 250 mls (8 ounces) Skin Contact May cause irritation with redness and pain May be absorbed through the skin with possible systemic effects Eye Contact Vapours cause eye irritation Splashes cause severe irritation possible corneal burns and eye damage Chronic Exposure Chronic exposure may cause skin effects Aggravation of Pre-existing Conditions Persons with pre-existing skin disorders or impaired liver kidney or pulmonary function may be more susceptible to the effects of this agent
13
METHYLENE BLUE ndash MSDS HAND OUTS CHEMWATCH ISSUE DATE 15052010 METHYLENE BLUE SAFETY-DATA(tm) Ratings (Provided here for your convenience)
----------------------------------------------------------------------------------------------------------- Health Rating 2 - Moderate Flammability Rating 1 - Slight Reactivity Rating 1 - Slight Contact Rating 1 - Slight Lab Protective Equip GOGGLES LAB COAT VENT HOOD PROPER GLOVES Storage Colour Code Green (General Storage) -----------------------------------------------------------------------------------------------------------
Potential Health Effects ----------------------------------
This material is relatively nonhazardous in routine industrial situations
Inhalation No adverse health effects expected from inhalation May cause a short period of rapid or difficult breathing Ingestion A burning sensation of the mouth may be noted following ingestion of methylene blue May cause nausea vomiting diarrhea and gastritis Large doses may cause abdominal and chest pain headache profuse sweating mental confusion painful maturation and methemoglobinemia Skin Contact Not expected to be a health hazard from skin exposure Methylene blue may colour the skin a bluish colour May cause photosensitization Eye Contact No adverse effects expected May cause mechanical irritation Chronic Exposure No information found Aggravation of Pre-existing Conditions No information found
14
NEW WORDS TO WORD BANKADD THESE TO YOUR EXCEL ALPHA TABLE
Precipitate Spooling Denaturising Lysing Homogenized Aggregate Lipid Centrifuging PCR
WORDS MEANING
15
bullDO YOU THINK YOU HAVE VERY MUCH IN COMMON WITH A KIWI FRUIT
BELIEVE IT OR NOT A KIWIS GENETIC MATERIAL IS VERY SIMILAR TO YOUR OWN
SEE AND TOUCH THE GENETIC MATERIAL THAT YOULL EXTRACT FROM THE CELLS OF A KIWI FRUIT
Activity Overview
Extract DNA from kiwi fruit using simple household chemicals
16
HT
TP
LEA
RN
GEN
ETIC
SU
TA
HE
DU
CO
NTE
NTL
AB
SE
XTR
AC
TIO
NH
OW
TO
DNA EXTRACTION USING KIWI FRUIT
17
AIM
TO
EX
TR
AC
T D
NA
FR
OM
TH
E C
ELLS
OF K
IWI F
RU
IT
wwwexploratoriumedu
MATERIALS EQUIPMENT (per student group takes 30 minutes)
frac12 cup Kiwi fruit 200 mL water dishwashing detergent dropping pipette fine mesh kitchen strainer amp cheese cloth glass rod large beaker large test tube light microscope meat tenderiser methylene blue microscope lamp inoculation needle microscope slide and cover slip paper towelling small beaker of alcohol (Isopropanol) spatula test-tube rack mortar and pestle
18
PART A EXTRACTING THE DNA
METHOD
1 Peal and Place the kiwi and water in the beaker and mush it with a mortar
and pestle DNA SOURCE About 125 ml 12 cup Twice as much water as DNA source (250
ml 1 cup) Table salt large pinch 1g 14 teaspoon Stir until the mixture is of a thin soupy
consistency
TAKE NOTES ndash NO TEXTBOOKS ALLOWED WHEN PERFORMING EXPERIMENT ndashMUST RECALL PROCEDURES METHOD
19
WHAT MUST I DO NOW
STRAIN THE DNA MIXTURE
DISHWASHING DETERGENT
20
2 Pour the thin cell mixture through the kitchen strainer (cheesecloth) into another large beaker
3 Measure 12 ml of the soup into
a small beakerAdd 2 ml of liquid detergentSwirl to mix stir thoroughly using a
glass rodLet mixture sit in container with
hot tap water (60 - 65degC) for 5 - 10 minutes
21
WHAT MUST I DO NOWMEAT TENDERIZER
ENZYME POWDERALCOHOL SEPARATION
wwwforumsoverclockerscomau
22
4 Add a pinch of enzyme (meat tenderizer) to mixture
(Using pineapple juice or contact lens cleaning solution will do the same as tenderizer)
Gently stir with toothpick skewer for 5 minutes continue stirring not too vigorously Be careful If you stir too hard youll break up the DNA making it harder to see
5 Quarter-fill a large test tube with the mixture
23
SLOWLY pour the same amount ice - cold (70 - 95 isopropyl or ethyl alcohol) into test tube pour alcohol down the side of the test tube
Tilting the test tube will make this easier to do
It forms a layer on top of the cell mixture
DO NOT MIX THE TWO LAYERS TOGETHER
Amount of alcohol and mixture should be the same
24
SEPARATION USING ALCOHOL
25
7 Observe the mixture for a few minutes You will see a white threadlike substance rise from the mixture to rest above in the alcohol layer
This is the DNA that you have extracted from the cells of the kiwi fruit
8 You can get more DNA to precipitate from the solution using a DNA collecting tool (glass or paper clip hook or cut inoculation needle)
Gently lift the water solution up into the alcohol layer (this allows more DNA to get in contact with the alcohol and precipitate)
26
INFORMATION DNA precipitates as a white stringy
ldquosnottyrdquo film at the water - alcohol interface and eventually will rise into the alcohol layer from the mixture layer
Allow test tube to sit for several minutes The clearer the DNA is the fewer
impurities you have
If you have an acceptable amount of DNA it can be spooled by rotating your collecting tool and then transferred into a clean tube container
27
WOW ndash SPOOLING THE DNA If you are careful you may be able wind up the DNA
around a glass rod or a skewer Position the tip of the glass rod or skewer where you can see the threads of DNA Steadily twist the rod or skewer as if you were making candy floss Alternatively use a straw to pull it out by suction ndash be careful not to get in you mouth
Donrsquot go too quickly
You should be able to pull the strands of DNA out of the mixture
ASK STUDENTS TO TAKE MOBILE PHONE PICTURES OF THEIR OWN EXTRACTED DNA AND COMPARE APPEARANCE WITH OTHERS
Photos of steps can be inserted in flow diagram(ICT)
28
KIWIDNA
29
If you want to save your DNA you can transfer it to a small container filled with alcohol
Leave tube container uncapped until the ethanol has evaporated
DNA can be stored in the fridge (dry or water buffer can be added)
You can now investigate the property
30
WHAT IS THAT STRINGY STUFFDNA IS A LONG STRINGY MOLECULE THE SALT THAT YOU ADDED IN STEP ONE HELPS IT STICK TOGETHER SO WHAT YOU SEE ARE CLUMPS OF TANGLED DNA MOLECULES
DNA NORMALLY STAYS DISSOLVED IN WATER BUT WHEN SALTY DNA COMES IN CONTACT WITH ALCOHOL IT BECOMES UNDISSOLVED THIS IS CALLED PRECIPITATION THE PHYSICAL FORCE OF THE DNA CLUMPING TOGETHER AS IT PRECIPITATES PULLS MORE STRANDS ALONG WITH IT AS IT RISES INTO THE ALCOHOL
31
THE WHYrsquoS Blending separated the pea cells In order to extract DNA from a cell the associated
membranes and proteins must first be removed (break apart the cells) and then physically separated (loosen the tough cell wall) from the DNA
To see the DNA we have to break open these two sacks We do this with detergent and salt( Sodium can be
involved in several of the steps ) Sodium is an element Its chemical symbol is Na for
Natrium the Latin word for sodium It is a positive ion and often associates with negative ions as part of useful compounds
Salt help precipitate protein and carbohydrates away from the DNA
Salt helps strip away the proteins associated with DNA + and ndash charge
32
THE WHYrsquoS Salt and Detergents are used to break down
cell walls and nuclear membranes to release the DNA
They work by chemically poking holes in the cell membranes or walls
Once holes are poked in the membranes the membranes can be further disrupted mechanically as with a blender
After that it is easier to get the contents of the cell out including the DNA
LETS HAVE A LOOK AT THE CELL
33
CELL TO DNA
httplearngeneticsutaheducontentlabsextractionhowtoHOW TO EXTRACT DNA FROM ANYTHING LIVING
34
THE WHYrsquoS WHY DETERGENT
Each cell is surrounded by a sack (the cell membrane)
DNA is found inside the second sack (nucleus) within each cell
To see the DNA we have to break it open A cell membrane has 2 layers of lipid(fat)
molecules with protein going through them When the lysis buffer (detergent) comes
close to the cell it captures the lipids and the proteins - breaks open the cell destroying the fatty membrane - DNA now released into the solution
35
A CELLS MEMBRANES HAVE TWO LAYERS OF LIPID (FAT) MOLECULES WITH PROTEINS GOING THROUGH THEM
36
Why detergent How does detergent work
Think about why you use soap to wash dishes or your hands To remove grease and dirt right
Soap molecules and grease molecules are made of two parts
1(Blue) Heads which like water 2(Green) Tails which hate water
37
AFTER ADDING THE DETERGENT WHAT DO YOU HAVE IN YOUR SOUP
38
THE WHYrsquoS ENZYME (MEAT TENDERIZER)
The DNA in the nucleus of the cell is molded folded and protected by proteins The tenderizer cut the proteins away from the DNA
In this experiment meat tenderizer acts as an enzyme to cut proteins just like a pair of scissors
The meat tenderizer cuts the proteins away from the DNA
39
THE WHYrsquoS CUTTING PROTEIN AND DNA
The DNA in the nucleus of the cell is moulded folded and protected by proteins
40
THE WHYrsquoSALCOHOL Alcohol is less dense than water so it floats
on top Look for clumps of white stringy stuff where the water and alcohol layers meet
DNA is not soluble in alcohol - other cell parts are
By adding alcohol DNA precipitates out of the solution and collect at the interface of the alcohol and soap layer
The colder the alcohol the less soluble the DNA will be in it
41
COLD ALCOHOL
DNA dissolves in water but precipitates in alcohol
Cold alcohol is used to separate DNA out of water-based solutions
This allows the DNA to be purified for subsequent genetic testing
Adding alcohol to a solution containing DNA is a simple way to obtain the pure DNA required and colder temperatures slow down enzymes that can break down DNA giving better extraction results
42
WHY IS DNA EXTRACTION IMPORTANT DNA extraction is an important molecular
biology procedure
By definition extraction is taking DNA out of any type of cell for the purpose of analysis
See Handouts for more information on the use of DNA extracted (extension only)
43
SUMMARIZEDNA EXTRACTING FROM KIWI FRUIT
44
BREAK DOWN CELL WALLS
Cells walls have to be destroyed to reach the DNA within cells
Extraction procedures to obtain pure DNA have to get rid of all the molecular and chemical components of the tissue from which the DNA is being extracted
First steps involve lysing or destroying the cell walls This can be done with a variety of chemical agents that are caustic to cell membranes but do not harm the DNA Cells can also be sonicated homogenized or ground up to destroy membranes
45
REMOVING Removing Lipids
Once the cell walls have been destroyed a detergent is added to get rid of the fats and oils that make up the cell membranes Detergents cause the fats and oils to dissolve into the solution
Remove ProteinsProteins and enzymes can be digested by
adding a protease to the solution Proteases break down proteins into small peptides and amino acids
46
PRECIPITATING AND PURIFYING Precipitate DNA
Adding cold alcohol to a solution will cause DNA to precipitate and aggregate The DNA can be collected by centrifuging the sample and pouring off the liquid layer The DNA should exist as a small pellet in the bottom of the centrifuge tube
Purify DNAThe DNA can be washed by re-suspending it
in a cold alcohol solution and re-centrifuging it several times to obtain a very pure DNA sample Typical alcohols used to precipitate DNA include ethanol and Isopropanol This process leaves a very pure sample for stringent DNA testing
47
PART B OBSERVING EXTRACTED DNA UNDER THE MICROSCOPE
48
PART B A CLOSER LOOK
8 Use a dropping pipette to carefully remove some of the threadlike substance from the top of your preparation
9 Place one or two drops onto the middle of a
microscope slide
10 Add two drops of methylene blue Wait 3 or 4 minutes to allow the methylene blue to be absorbed by the DNA
11 Carefully place a cover slip on the slide Gently press
a folded piece of paper towelling over the top of the prepared slide to soak up any excess liquid
12 Observe the DNA under low power then high power
49
DISCUSSIONANSWER IN GROUP RELATED WORKED 1 Write a detailed description of the material
floating at the top of the test tube after the alcohol was added
2 Describe the DNA as it appears under the
microscope under high power 3 Prepare a diagram of your DNA specimen 4 What was the reason for using methylene
blue
50
HOMEWORKCONSTRUCT A FLOW CHART DRAWING MIND MAP OF THE PROCESS YOU USED EXTRACTING THE DNA FROM KIWIrsquoS
Next to each step explain how the method you used was important (Hint What substances make up the membranes of cells and cell organelles
How do detergents work
What is the active ingredient in meat tenderiser
51
TIPS STEPS FOR FLOW CHART In order to release the DNA from the
nuclei of the pea cells you first separated the cells from one another
Then the cell and nuclear membranes needed to be ruptured to release the cell contents and the contents of the nucleus
Once removed from the nuclear membrane the DNA had to be untangled into the visible threadlike structures you ended up with
52
HOMEWORK EXTENSION GROUP QUESTIONS Where can DNA be found in the cell Discuss the action of the soap (detergent) on the cell
What is the purpose of the soap in this activity What was the purpose of the Sodium Chloride Include a
discussion of polarity and charged particles Why was the cold ethanol added to the soap and salt
mixture Describe the appearance of your final product Draw a diagram of DNA containing 5 sets of nucleotide
bases labelling the hydrogen bonds between the bases References and Resources Adapted from Berry Full
of DNA by Diane Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
53
HTTPWWWGENOMEBCCAEDUCATIONTEACHERSCLASSROOM-ACTIVITIESDNA-EXTRACTION
54
EXTRACHROMOSOMES EXIST IN PAIRS Because our chromosomes exist in pairs (and consequently we
have 2 alleles of each gene) we are a diploid species This is why our somatic cells are represented as 2n Our gametes (sperm and ova) on the other hand are haploid and are represented as n Other species may have different ploidy for example
triploid (3n) seedless watermelons tetaploid (4n) salmonidae fish pentaploid (5n) Kenai birch hexaploid (6n) some types of wheat kiwi fruit octaploid (8n) acipenser (a genus
of sturgeon fish strawberies) decaploid (10n) some strawberries dodecaploid (12n) some types of amphibians eg Xenopus
ruwenzoriensis
wwwcyberusca
55
YOUR EXTRACTED DNA UNDER THE MICROSCOPE LOOKED
LIKE
Collaboration-Brainstorming on what to look at under the
- DNA extraction- DNA is too small for even a microscope to see and after the extraction it just appears like a blob True or false
Could you any DNA strand
wwwemployeescsbsjuedu
56
HANDOUT ON DNA EXTRACTIONBACKGROUND EVERY DAY LIFE
EXTRA READING MATERIAL(EXTENSION )
DISCOVERING DNA DNA ON THE INSIDE USE OF DNA EXTRACTION INN EVERYDAY LIFE WHY IS DNA TESTING GOOD MOLECULAR BIOLOGY PCR-based diagnostics of genomic DNA
57
REFERENCES httpwwwsquidoocomhow-to-get-dna-from-a-kiwi-fruit Why Is DNA Testing Good | eHowcom
httpwwwehowcomabout_6684410_dna-testing-good_htmlixzz1MkbcIJs5 Why Is DNA Extraction Important | eHowcom
httpwwwehowcomlist_5839095_dna-extraction-important_htmlixzz1MkZDrqyY
Why Is Sodium Used in DNA Extraction | eHowcom httpwwwehowcomabout_6504902_sodium-used-dna-extraction_htmlixzz1MkaGWg57
Why Is Cold Alcohol Used in DNA Tests | eHowcom httpwwwehowcomabout_6399349_cold-alcohol-used-dna-tests_htmlixzz1MkbEmEdu
httplearngeneticsutaheducontentlabsextractionhowto www kamibudaksainsblogspotcom
httpwwwchemwatch goldcom References and Resources Adapted from Berry Full of DNA by Diane
Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
58
REFERENCES
Uses of DNA Extraction | eHowcom httpwwwehowcomabout_5344428_uses-dna-extractionhtmlixzz1MkWQB2TZ
Science 10 A Contextual Approach Heineman Queensland Science Projects Regan Spence Maggie Spenceley
httpenwikipediaorgwikiMolecular_biology httpwwwclinchemorgcgicontent
extract44102201 http
employeescsbsjueduhjakubowskiclassesch331dnaoldnastructurehtml
httpwwwgenomebccaeducationteachersclassroom-activitiesdna-extraction
7
FIGURE HTTPEMPLOYEESCSBSJUEDUHJAKUBOWSKICLASSESCH331DNAOLDNASTRUCTUREHTML
PACKAGING OF DNA IN THE NUCLEUS
8
httpwwwyoutubecomwatchv=tmVP1cotozc
9
DNA extraction refers to the process that scientists use to break down a piece of evidence such as a piece of hair or a drop of blood in order to determine the individuals DNA which is unique to each person Understand the process of DNA extraction with information from a biology teacher in this free video on science
Expert Janice CrenettiContact WeAreHDTVcomBio Janice Creneti has a Bachelor of Science in secondary science education and a Bachelor of Art in biology from Boston UniversityFilmmaker Christopher Rokosz
Category Entertainment Tags science dna dna testing dna structure dna model dna replication dna molecule
10
REVIEWING OF INFORMATION ONLABORATORY SAFETY RULES AND RISK ASSESSMENTS
Do not enter the laboratory unless you are with a teacher Never touch equipment in the laboratory unless you are told to use it Donrsquot eat or drink in the laboratory Always walkmdashnever run Wear protective clothingmdash a laboratory coat or apron and when
appropriate safety glasses Never taste anything Tie up long hair Always point test tubes away from people Check with your teacher on how to dispose of waste liquids and solids
Broken glass should be cleaned up using gloves a brush and dustpan and placed in a special bin
If you spill something on your skin or clothes wash it immediately with lots of water Tell your teacher
Report all accidents and breakages to your teacher After heating equipment let it cool on a heatproof mat before picking
it up This will avoid burns Clean all equipment after use and put it back where you got it from
Clean and dry your work bench
11
RISK ASSESSMENT ON EXPERIMENT AND INVESTIGATION ndash KIWI FRUIT
Eyewear and protective clothingNo eat or drinkingCover the MSDSrsquo e on Isopropanol and Methylene blue ndash hard copy handouts (health and safety regulation)
Any broken glassware must be reported
Care must be taken when using and washing the blender ndash sharp blades
12
ISOPROPANOL ndash MSDS HAND OUTS CHEMWATCH ISSUE DATE 15052010 ISOPROPANOL - FLAMMABLE Dangerous Good 3 SAFETY-DATA(tm) Ratings (Provided here for your convenience)
----------------------------------------------------------------------------------------------------------- Health Rating 2 - Moderate Flammability Rating 3 - Severe (Flammable) Reactivity Rating 2 - Moderate Contact Rating 3 - Severe Lab Protective Equip GOGGLES amp SHIELD LAB COAT amp APRON VENT HOOD PROPER GLOVES CLASS B EXTINGUISHER Storage Colour Code Red (Flammable)
Potential Health Effects ----------------------------------
Inhalation Inhalation of vapours irritates the respiratory tract Exposure to high concentrations has a narcotic effect producing symptoms of dizziness drowsiness headache staggering unconsciousness and possibly death Ingestion Can cause drowsiness unconsciousness and death Gastrointestinal pain cramps nausea vomiting and diarrhoea may also result The single lethal dose for a human adult = about 250 mls (8 ounces) Skin Contact May cause irritation with redness and pain May be absorbed through the skin with possible systemic effects Eye Contact Vapours cause eye irritation Splashes cause severe irritation possible corneal burns and eye damage Chronic Exposure Chronic exposure may cause skin effects Aggravation of Pre-existing Conditions Persons with pre-existing skin disorders or impaired liver kidney or pulmonary function may be more susceptible to the effects of this agent
13
METHYLENE BLUE ndash MSDS HAND OUTS CHEMWATCH ISSUE DATE 15052010 METHYLENE BLUE SAFETY-DATA(tm) Ratings (Provided here for your convenience)
----------------------------------------------------------------------------------------------------------- Health Rating 2 - Moderate Flammability Rating 1 - Slight Reactivity Rating 1 - Slight Contact Rating 1 - Slight Lab Protective Equip GOGGLES LAB COAT VENT HOOD PROPER GLOVES Storage Colour Code Green (General Storage) -----------------------------------------------------------------------------------------------------------
Potential Health Effects ----------------------------------
This material is relatively nonhazardous in routine industrial situations
Inhalation No adverse health effects expected from inhalation May cause a short period of rapid or difficult breathing Ingestion A burning sensation of the mouth may be noted following ingestion of methylene blue May cause nausea vomiting diarrhea and gastritis Large doses may cause abdominal and chest pain headache profuse sweating mental confusion painful maturation and methemoglobinemia Skin Contact Not expected to be a health hazard from skin exposure Methylene blue may colour the skin a bluish colour May cause photosensitization Eye Contact No adverse effects expected May cause mechanical irritation Chronic Exposure No information found Aggravation of Pre-existing Conditions No information found
14
NEW WORDS TO WORD BANKADD THESE TO YOUR EXCEL ALPHA TABLE
Precipitate Spooling Denaturising Lysing Homogenized Aggregate Lipid Centrifuging PCR
WORDS MEANING
15
bullDO YOU THINK YOU HAVE VERY MUCH IN COMMON WITH A KIWI FRUIT
BELIEVE IT OR NOT A KIWIS GENETIC MATERIAL IS VERY SIMILAR TO YOUR OWN
SEE AND TOUCH THE GENETIC MATERIAL THAT YOULL EXTRACT FROM THE CELLS OF A KIWI FRUIT
Activity Overview
Extract DNA from kiwi fruit using simple household chemicals
16
HT
TP
LEA
RN
GEN
ETIC
SU
TA
HE
DU
CO
NTE
NTL
AB
SE
XTR
AC
TIO
NH
OW
TO
DNA EXTRACTION USING KIWI FRUIT
17
AIM
TO
EX
TR
AC
T D
NA
FR
OM
TH
E C
ELLS
OF K
IWI F
RU
IT
wwwexploratoriumedu
MATERIALS EQUIPMENT (per student group takes 30 minutes)
frac12 cup Kiwi fruit 200 mL water dishwashing detergent dropping pipette fine mesh kitchen strainer amp cheese cloth glass rod large beaker large test tube light microscope meat tenderiser methylene blue microscope lamp inoculation needle microscope slide and cover slip paper towelling small beaker of alcohol (Isopropanol) spatula test-tube rack mortar and pestle
18
PART A EXTRACTING THE DNA
METHOD
1 Peal and Place the kiwi and water in the beaker and mush it with a mortar
and pestle DNA SOURCE About 125 ml 12 cup Twice as much water as DNA source (250
ml 1 cup) Table salt large pinch 1g 14 teaspoon Stir until the mixture is of a thin soupy
consistency
TAKE NOTES ndash NO TEXTBOOKS ALLOWED WHEN PERFORMING EXPERIMENT ndashMUST RECALL PROCEDURES METHOD
19
WHAT MUST I DO NOW
STRAIN THE DNA MIXTURE
DISHWASHING DETERGENT
20
2 Pour the thin cell mixture through the kitchen strainer (cheesecloth) into another large beaker
3 Measure 12 ml of the soup into
a small beakerAdd 2 ml of liquid detergentSwirl to mix stir thoroughly using a
glass rodLet mixture sit in container with
hot tap water (60 - 65degC) for 5 - 10 minutes
21
WHAT MUST I DO NOWMEAT TENDERIZER
ENZYME POWDERALCOHOL SEPARATION
wwwforumsoverclockerscomau
22
4 Add a pinch of enzyme (meat tenderizer) to mixture
(Using pineapple juice or contact lens cleaning solution will do the same as tenderizer)
Gently stir with toothpick skewer for 5 minutes continue stirring not too vigorously Be careful If you stir too hard youll break up the DNA making it harder to see
5 Quarter-fill a large test tube with the mixture
23
SLOWLY pour the same amount ice - cold (70 - 95 isopropyl or ethyl alcohol) into test tube pour alcohol down the side of the test tube
Tilting the test tube will make this easier to do
It forms a layer on top of the cell mixture
DO NOT MIX THE TWO LAYERS TOGETHER
Amount of alcohol and mixture should be the same
24
SEPARATION USING ALCOHOL
25
7 Observe the mixture for a few minutes You will see a white threadlike substance rise from the mixture to rest above in the alcohol layer
This is the DNA that you have extracted from the cells of the kiwi fruit
8 You can get more DNA to precipitate from the solution using a DNA collecting tool (glass or paper clip hook or cut inoculation needle)
Gently lift the water solution up into the alcohol layer (this allows more DNA to get in contact with the alcohol and precipitate)
26
INFORMATION DNA precipitates as a white stringy
ldquosnottyrdquo film at the water - alcohol interface and eventually will rise into the alcohol layer from the mixture layer
Allow test tube to sit for several minutes The clearer the DNA is the fewer
impurities you have
If you have an acceptable amount of DNA it can be spooled by rotating your collecting tool and then transferred into a clean tube container
27
WOW ndash SPOOLING THE DNA If you are careful you may be able wind up the DNA
around a glass rod or a skewer Position the tip of the glass rod or skewer where you can see the threads of DNA Steadily twist the rod or skewer as if you were making candy floss Alternatively use a straw to pull it out by suction ndash be careful not to get in you mouth
Donrsquot go too quickly
You should be able to pull the strands of DNA out of the mixture
ASK STUDENTS TO TAKE MOBILE PHONE PICTURES OF THEIR OWN EXTRACTED DNA AND COMPARE APPEARANCE WITH OTHERS
Photos of steps can be inserted in flow diagram(ICT)
28
KIWIDNA
29
If you want to save your DNA you can transfer it to a small container filled with alcohol
Leave tube container uncapped until the ethanol has evaporated
DNA can be stored in the fridge (dry or water buffer can be added)
You can now investigate the property
30
WHAT IS THAT STRINGY STUFFDNA IS A LONG STRINGY MOLECULE THE SALT THAT YOU ADDED IN STEP ONE HELPS IT STICK TOGETHER SO WHAT YOU SEE ARE CLUMPS OF TANGLED DNA MOLECULES
DNA NORMALLY STAYS DISSOLVED IN WATER BUT WHEN SALTY DNA COMES IN CONTACT WITH ALCOHOL IT BECOMES UNDISSOLVED THIS IS CALLED PRECIPITATION THE PHYSICAL FORCE OF THE DNA CLUMPING TOGETHER AS IT PRECIPITATES PULLS MORE STRANDS ALONG WITH IT AS IT RISES INTO THE ALCOHOL
31
THE WHYrsquoS Blending separated the pea cells In order to extract DNA from a cell the associated
membranes and proteins must first be removed (break apart the cells) and then physically separated (loosen the tough cell wall) from the DNA
To see the DNA we have to break open these two sacks We do this with detergent and salt( Sodium can be
involved in several of the steps ) Sodium is an element Its chemical symbol is Na for
Natrium the Latin word for sodium It is a positive ion and often associates with negative ions as part of useful compounds
Salt help precipitate protein and carbohydrates away from the DNA
Salt helps strip away the proteins associated with DNA + and ndash charge
32
THE WHYrsquoS Salt and Detergents are used to break down
cell walls and nuclear membranes to release the DNA
They work by chemically poking holes in the cell membranes or walls
Once holes are poked in the membranes the membranes can be further disrupted mechanically as with a blender
After that it is easier to get the contents of the cell out including the DNA
LETS HAVE A LOOK AT THE CELL
33
CELL TO DNA
httplearngeneticsutaheducontentlabsextractionhowtoHOW TO EXTRACT DNA FROM ANYTHING LIVING
34
THE WHYrsquoS WHY DETERGENT
Each cell is surrounded by a sack (the cell membrane)
DNA is found inside the second sack (nucleus) within each cell
To see the DNA we have to break it open A cell membrane has 2 layers of lipid(fat)
molecules with protein going through them When the lysis buffer (detergent) comes
close to the cell it captures the lipids and the proteins - breaks open the cell destroying the fatty membrane - DNA now released into the solution
35
A CELLS MEMBRANES HAVE TWO LAYERS OF LIPID (FAT) MOLECULES WITH PROTEINS GOING THROUGH THEM
36
Why detergent How does detergent work
Think about why you use soap to wash dishes or your hands To remove grease and dirt right
Soap molecules and grease molecules are made of two parts
1(Blue) Heads which like water 2(Green) Tails which hate water
37
AFTER ADDING THE DETERGENT WHAT DO YOU HAVE IN YOUR SOUP
38
THE WHYrsquoS ENZYME (MEAT TENDERIZER)
The DNA in the nucleus of the cell is molded folded and protected by proteins The tenderizer cut the proteins away from the DNA
In this experiment meat tenderizer acts as an enzyme to cut proteins just like a pair of scissors
The meat tenderizer cuts the proteins away from the DNA
39
THE WHYrsquoS CUTTING PROTEIN AND DNA
The DNA in the nucleus of the cell is moulded folded and protected by proteins
40
THE WHYrsquoSALCOHOL Alcohol is less dense than water so it floats
on top Look for clumps of white stringy stuff where the water and alcohol layers meet
DNA is not soluble in alcohol - other cell parts are
By adding alcohol DNA precipitates out of the solution and collect at the interface of the alcohol and soap layer
The colder the alcohol the less soluble the DNA will be in it
41
COLD ALCOHOL
DNA dissolves in water but precipitates in alcohol
Cold alcohol is used to separate DNA out of water-based solutions
This allows the DNA to be purified for subsequent genetic testing
Adding alcohol to a solution containing DNA is a simple way to obtain the pure DNA required and colder temperatures slow down enzymes that can break down DNA giving better extraction results
42
WHY IS DNA EXTRACTION IMPORTANT DNA extraction is an important molecular
biology procedure
By definition extraction is taking DNA out of any type of cell for the purpose of analysis
See Handouts for more information on the use of DNA extracted (extension only)
43
SUMMARIZEDNA EXTRACTING FROM KIWI FRUIT
44
BREAK DOWN CELL WALLS
Cells walls have to be destroyed to reach the DNA within cells
Extraction procedures to obtain pure DNA have to get rid of all the molecular and chemical components of the tissue from which the DNA is being extracted
First steps involve lysing or destroying the cell walls This can be done with a variety of chemical agents that are caustic to cell membranes but do not harm the DNA Cells can also be sonicated homogenized or ground up to destroy membranes
45
REMOVING Removing Lipids
Once the cell walls have been destroyed a detergent is added to get rid of the fats and oils that make up the cell membranes Detergents cause the fats and oils to dissolve into the solution
Remove ProteinsProteins and enzymes can be digested by
adding a protease to the solution Proteases break down proteins into small peptides and amino acids
46
PRECIPITATING AND PURIFYING Precipitate DNA
Adding cold alcohol to a solution will cause DNA to precipitate and aggregate The DNA can be collected by centrifuging the sample and pouring off the liquid layer The DNA should exist as a small pellet in the bottom of the centrifuge tube
Purify DNAThe DNA can be washed by re-suspending it
in a cold alcohol solution and re-centrifuging it several times to obtain a very pure DNA sample Typical alcohols used to precipitate DNA include ethanol and Isopropanol This process leaves a very pure sample for stringent DNA testing
47
PART B OBSERVING EXTRACTED DNA UNDER THE MICROSCOPE
48
PART B A CLOSER LOOK
8 Use a dropping pipette to carefully remove some of the threadlike substance from the top of your preparation
9 Place one or two drops onto the middle of a
microscope slide
10 Add two drops of methylene blue Wait 3 or 4 minutes to allow the methylene blue to be absorbed by the DNA
11 Carefully place a cover slip on the slide Gently press
a folded piece of paper towelling over the top of the prepared slide to soak up any excess liquid
12 Observe the DNA under low power then high power
49
DISCUSSIONANSWER IN GROUP RELATED WORKED 1 Write a detailed description of the material
floating at the top of the test tube after the alcohol was added
2 Describe the DNA as it appears under the
microscope under high power 3 Prepare a diagram of your DNA specimen 4 What was the reason for using methylene
blue
50
HOMEWORKCONSTRUCT A FLOW CHART DRAWING MIND MAP OF THE PROCESS YOU USED EXTRACTING THE DNA FROM KIWIrsquoS
Next to each step explain how the method you used was important (Hint What substances make up the membranes of cells and cell organelles
How do detergents work
What is the active ingredient in meat tenderiser
51
TIPS STEPS FOR FLOW CHART In order to release the DNA from the
nuclei of the pea cells you first separated the cells from one another
Then the cell and nuclear membranes needed to be ruptured to release the cell contents and the contents of the nucleus
Once removed from the nuclear membrane the DNA had to be untangled into the visible threadlike structures you ended up with
52
HOMEWORK EXTENSION GROUP QUESTIONS Where can DNA be found in the cell Discuss the action of the soap (detergent) on the cell
What is the purpose of the soap in this activity What was the purpose of the Sodium Chloride Include a
discussion of polarity and charged particles Why was the cold ethanol added to the soap and salt
mixture Describe the appearance of your final product Draw a diagram of DNA containing 5 sets of nucleotide
bases labelling the hydrogen bonds between the bases References and Resources Adapted from Berry Full
of DNA by Diane Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
53
HTTPWWWGENOMEBCCAEDUCATIONTEACHERSCLASSROOM-ACTIVITIESDNA-EXTRACTION
54
EXTRACHROMOSOMES EXIST IN PAIRS Because our chromosomes exist in pairs (and consequently we
have 2 alleles of each gene) we are a diploid species This is why our somatic cells are represented as 2n Our gametes (sperm and ova) on the other hand are haploid and are represented as n Other species may have different ploidy for example
triploid (3n) seedless watermelons tetaploid (4n) salmonidae fish pentaploid (5n) Kenai birch hexaploid (6n) some types of wheat kiwi fruit octaploid (8n) acipenser (a genus
of sturgeon fish strawberies) decaploid (10n) some strawberries dodecaploid (12n) some types of amphibians eg Xenopus
ruwenzoriensis
wwwcyberusca
55
YOUR EXTRACTED DNA UNDER THE MICROSCOPE LOOKED
LIKE
Collaboration-Brainstorming on what to look at under the
- DNA extraction- DNA is too small for even a microscope to see and after the extraction it just appears like a blob True or false
Could you any DNA strand
wwwemployeescsbsjuedu
56
HANDOUT ON DNA EXTRACTIONBACKGROUND EVERY DAY LIFE
EXTRA READING MATERIAL(EXTENSION )
DISCOVERING DNA DNA ON THE INSIDE USE OF DNA EXTRACTION INN EVERYDAY LIFE WHY IS DNA TESTING GOOD MOLECULAR BIOLOGY PCR-based diagnostics of genomic DNA
57
REFERENCES httpwwwsquidoocomhow-to-get-dna-from-a-kiwi-fruit Why Is DNA Testing Good | eHowcom
httpwwwehowcomabout_6684410_dna-testing-good_htmlixzz1MkbcIJs5 Why Is DNA Extraction Important | eHowcom
httpwwwehowcomlist_5839095_dna-extraction-important_htmlixzz1MkZDrqyY
Why Is Sodium Used in DNA Extraction | eHowcom httpwwwehowcomabout_6504902_sodium-used-dna-extraction_htmlixzz1MkaGWg57
Why Is Cold Alcohol Used in DNA Tests | eHowcom httpwwwehowcomabout_6399349_cold-alcohol-used-dna-tests_htmlixzz1MkbEmEdu
httplearngeneticsutaheducontentlabsextractionhowto www kamibudaksainsblogspotcom
httpwwwchemwatch goldcom References and Resources Adapted from Berry Full of DNA by Diane
Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
58
REFERENCES
Uses of DNA Extraction | eHowcom httpwwwehowcomabout_5344428_uses-dna-extractionhtmlixzz1MkWQB2TZ
Science 10 A Contextual Approach Heineman Queensland Science Projects Regan Spence Maggie Spenceley
httpenwikipediaorgwikiMolecular_biology httpwwwclinchemorgcgicontent
extract44102201 http
employeescsbsjueduhjakubowskiclassesch331dnaoldnastructurehtml
httpwwwgenomebccaeducationteachersclassroom-activitiesdna-extraction
8
httpwwwyoutubecomwatchv=tmVP1cotozc
9
DNA extraction refers to the process that scientists use to break down a piece of evidence such as a piece of hair or a drop of blood in order to determine the individuals DNA which is unique to each person Understand the process of DNA extraction with information from a biology teacher in this free video on science
Expert Janice CrenettiContact WeAreHDTVcomBio Janice Creneti has a Bachelor of Science in secondary science education and a Bachelor of Art in biology from Boston UniversityFilmmaker Christopher Rokosz
Category Entertainment Tags science dna dna testing dna structure dna model dna replication dna molecule
10
REVIEWING OF INFORMATION ONLABORATORY SAFETY RULES AND RISK ASSESSMENTS
Do not enter the laboratory unless you are with a teacher Never touch equipment in the laboratory unless you are told to use it Donrsquot eat or drink in the laboratory Always walkmdashnever run Wear protective clothingmdash a laboratory coat or apron and when
appropriate safety glasses Never taste anything Tie up long hair Always point test tubes away from people Check with your teacher on how to dispose of waste liquids and solids
Broken glass should be cleaned up using gloves a brush and dustpan and placed in a special bin
If you spill something on your skin or clothes wash it immediately with lots of water Tell your teacher
Report all accidents and breakages to your teacher After heating equipment let it cool on a heatproof mat before picking
it up This will avoid burns Clean all equipment after use and put it back where you got it from
Clean and dry your work bench
11
RISK ASSESSMENT ON EXPERIMENT AND INVESTIGATION ndash KIWI FRUIT
Eyewear and protective clothingNo eat or drinkingCover the MSDSrsquo e on Isopropanol and Methylene blue ndash hard copy handouts (health and safety regulation)
Any broken glassware must be reported
Care must be taken when using and washing the blender ndash sharp blades
12
ISOPROPANOL ndash MSDS HAND OUTS CHEMWATCH ISSUE DATE 15052010 ISOPROPANOL - FLAMMABLE Dangerous Good 3 SAFETY-DATA(tm) Ratings (Provided here for your convenience)
----------------------------------------------------------------------------------------------------------- Health Rating 2 - Moderate Flammability Rating 3 - Severe (Flammable) Reactivity Rating 2 - Moderate Contact Rating 3 - Severe Lab Protective Equip GOGGLES amp SHIELD LAB COAT amp APRON VENT HOOD PROPER GLOVES CLASS B EXTINGUISHER Storage Colour Code Red (Flammable)
Potential Health Effects ----------------------------------
Inhalation Inhalation of vapours irritates the respiratory tract Exposure to high concentrations has a narcotic effect producing symptoms of dizziness drowsiness headache staggering unconsciousness and possibly death Ingestion Can cause drowsiness unconsciousness and death Gastrointestinal pain cramps nausea vomiting and diarrhoea may also result The single lethal dose for a human adult = about 250 mls (8 ounces) Skin Contact May cause irritation with redness and pain May be absorbed through the skin with possible systemic effects Eye Contact Vapours cause eye irritation Splashes cause severe irritation possible corneal burns and eye damage Chronic Exposure Chronic exposure may cause skin effects Aggravation of Pre-existing Conditions Persons with pre-existing skin disorders or impaired liver kidney or pulmonary function may be more susceptible to the effects of this agent
13
METHYLENE BLUE ndash MSDS HAND OUTS CHEMWATCH ISSUE DATE 15052010 METHYLENE BLUE SAFETY-DATA(tm) Ratings (Provided here for your convenience)
----------------------------------------------------------------------------------------------------------- Health Rating 2 - Moderate Flammability Rating 1 - Slight Reactivity Rating 1 - Slight Contact Rating 1 - Slight Lab Protective Equip GOGGLES LAB COAT VENT HOOD PROPER GLOVES Storage Colour Code Green (General Storage) -----------------------------------------------------------------------------------------------------------
Potential Health Effects ----------------------------------
This material is relatively nonhazardous in routine industrial situations
Inhalation No adverse health effects expected from inhalation May cause a short period of rapid or difficult breathing Ingestion A burning sensation of the mouth may be noted following ingestion of methylene blue May cause nausea vomiting diarrhea and gastritis Large doses may cause abdominal and chest pain headache profuse sweating mental confusion painful maturation and methemoglobinemia Skin Contact Not expected to be a health hazard from skin exposure Methylene blue may colour the skin a bluish colour May cause photosensitization Eye Contact No adverse effects expected May cause mechanical irritation Chronic Exposure No information found Aggravation of Pre-existing Conditions No information found
14
NEW WORDS TO WORD BANKADD THESE TO YOUR EXCEL ALPHA TABLE
Precipitate Spooling Denaturising Lysing Homogenized Aggregate Lipid Centrifuging PCR
WORDS MEANING
15
bullDO YOU THINK YOU HAVE VERY MUCH IN COMMON WITH A KIWI FRUIT
BELIEVE IT OR NOT A KIWIS GENETIC MATERIAL IS VERY SIMILAR TO YOUR OWN
SEE AND TOUCH THE GENETIC MATERIAL THAT YOULL EXTRACT FROM THE CELLS OF A KIWI FRUIT
Activity Overview
Extract DNA from kiwi fruit using simple household chemicals
16
HT
TP
LEA
RN
GEN
ETIC
SU
TA
HE
DU
CO
NTE
NTL
AB
SE
XTR
AC
TIO
NH
OW
TO
DNA EXTRACTION USING KIWI FRUIT
17
AIM
TO
EX
TR
AC
T D
NA
FR
OM
TH
E C
ELLS
OF K
IWI F
RU
IT
wwwexploratoriumedu
MATERIALS EQUIPMENT (per student group takes 30 minutes)
frac12 cup Kiwi fruit 200 mL water dishwashing detergent dropping pipette fine mesh kitchen strainer amp cheese cloth glass rod large beaker large test tube light microscope meat tenderiser methylene blue microscope lamp inoculation needle microscope slide and cover slip paper towelling small beaker of alcohol (Isopropanol) spatula test-tube rack mortar and pestle
18
PART A EXTRACTING THE DNA
METHOD
1 Peal and Place the kiwi and water in the beaker and mush it with a mortar
and pestle DNA SOURCE About 125 ml 12 cup Twice as much water as DNA source (250
ml 1 cup) Table salt large pinch 1g 14 teaspoon Stir until the mixture is of a thin soupy
consistency
TAKE NOTES ndash NO TEXTBOOKS ALLOWED WHEN PERFORMING EXPERIMENT ndashMUST RECALL PROCEDURES METHOD
19
WHAT MUST I DO NOW
STRAIN THE DNA MIXTURE
DISHWASHING DETERGENT
20
2 Pour the thin cell mixture through the kitchen strainer (cheesecloth) into another large beaker
3 Measure 12 ml of the soup into
a small beakerAdd 2 ml of liquid detergentSwirl to mix stir thoroughly using a
glass rodLet mixture sit in container with
hot tap water (60 - 65degC) for 5 - 10 minutes
21
WHAT MUST I DO NOWMEAT TENDERIZER
ENZYME POWDERALCOHOL SEPARATION
wwwforumsoverclockerscomau
22
4 Add a pinch of enzyme (meat tenderizer) to mixture
(Using pineapple juice or contact lens cleaning solution will do the same as tenderizer)
Gently stir with toothpick skewer for 5 minutes continue stirring not too vigorously Be careful If you stir too hard youll break up the DNA making it harder to see
5 Quarter-fill a large test tube with the mixture
23
SLOWLY pour the same amount ice - cold (70 - 95 isopropyl or ethyl alcohol) into test tube pour alcohol down the side of the test tube
Tilting the test tube will make this easier to do
It forms a layer on top of the cell mixture
DO NOT MIX THE TWO LAYERS TOGETHER
Amount of alcohol and mixture should be the same
24
SEPARATION USING ALCOHOL
25
7 Observe the mixture for a few minutes You will see a white threadlike substance rise from the mixture to rest above in the alcohol layer
This is the DNA that you have extracted from the cells of the kiwi fruit
8 You can get more DNA to precipitate from the solution using a DNA collecting tool (glass or paper clip hook or cut inoculation needle)
Gently lift the water solution up into the alcohol layer (this allows more DNA to get in contact with the alcohol and precipitate)
26
INFORMATION DNA precipitates as a white stringy
ldquosnottyrdquo film at the water - alcohol interface and eventually will rise into the alcohol layer from the mixture layer
Allow test tube to sit for several minutes The clearer the DNA is the fewer
impurities you have
If you have an acceptable amount of DNA it can be spooled by rotating your collecting tool and then transferred into a clean tube container
27
WOW ndash SPOOLING THE DNA If you are careful you may be able wind up the DNA
around a glass rod or a skewer Position the tip of the glass rod or skewer where you can see the threads of DNA Steadily twist the rod or skewer as if you were making candy floss Alternatively use a straw to pull it out by suction ndash be careful not to get in you mouth
Donrsquot go too quickly
You should be able to pull the strands of DNA out of the mixture
ASK STUDENTS TO TAKE MOBILE PHONE PICTURES OF THEIR OWN EXTRACTED DNA AND COMPARE APPEARANCE WITH OTHERS
Photos of steps can be inserted in flow diagram(ICT)
28
KIWIDNA
29
If you want to save your DNA you can transfer it to a small container filled with alcohol
Leave tube container uncapped until the ethanol has evaporated
DNA can be stored in the fridge (dry or water buffer can be added)
You can now investigate the property
30
WHAT IS THAT STRINGY STUFFDNA IS A LONG STRINGY MOLECULE THE SALT THAT YOU ADDED IN STEP ONE HELPS IT STICK TOGETHER SO WHAT YOU SEE ARE CLUMPS OF TANGLED DNA MOLECULES
DNA NORMALLY STAYS DISSOLVED IN WATER BUT WHEN SALTY DNA COMES IN CONTACT WITH ALCOHOL IT BECOMES UNDISSOLVED THIS IS CALLED PRECIPITATION THE PHYSICAL FORCE OF THE DNA CLUMPING TOGETHER AS IT PRECIPITATES PULLS MORE STRANDS ALONG WITH IT AS IT RISES INTO THE ALCOHOL
31
THE WHYrsquoS Blending separated the pea cells In order to extract DNA from a cell the associated
membranes and proteins must first be removed (break apart the cells) and then physically separated (loosen the tough cell wall) from the DNA
To see the DNA we have to break open these two sacks We do this with detergent and salt( Sodium can be
involved in several of the steps ) Sodium is an element Its chemical symbol is Na for
Natrium the Latin word for sodium It is a positive ion and often associates with negative ions as part of useful compounds
Salt help precipitate protein and carbohydrates away from the DNA
Salt helps strip away the proteins associated with DNA + and ndash charge
32
THE WHYrsquoS Salt and Detergents are used to break down
cell walls and nuclear membranes to release the DNA
They work by chemically poking holes in the cell membranes or walls
Once holes are poked in the membranes the membranes can be further disrupted mechanically as with a blender
After that it is easier to get the contents of the cell out including the DNA
LETS HAVE A LOOK AT THE CELL
33
CELL TO DNA
httplearngeneticsutaheducontentlabsextractionhowtoHOW TO EXTRACT DNA FROM ANYTHING LIVING
34
THE WHYrsquoS WHY DETERGENT
Each cell is surrounded by a sack (the cell membrane)
DNA is found inside the second sack (nucleus) within each cell
To see the DNA we have to break it open A cell membrane has 2 layers of lipid(fat)
molecules with protein going through them When the lysis buffer (detergent) comes
close to the cell it captures the lipids and the proteins - breaks open the cell destroying the fatty membrane - DNA now released into the solution
35
A CELLS MEMBRANES HAVE TWO LAYERS OF LIPID (FAT) MOLECULES WITH PROTEINS GOING THROUGH THEM
36
Why detergent How does detergent work
Think about why you use soap to wash dishes or your hands To remove grease and dirt right
Soap molecules and grease molecules are made of two parts
1(Blue) Heads which like water 2(Green) Tails which hate water
37
AFTER ADDING THE DETERGENT WHAT DO YOU HAVE IN YOUR SOUP
38
THE WHYrsquoS ENZYME (MEAT TENDERIZER)
The DNA in the nucleus of the cell is molded folded and protected by proteins The tenderizer cut the proteins away from the DNA
In this experiment meat tenderizer acts as an enzyme to cut proteins just like a pair of scissors
The meat tenderizer cuts the proteins away from the DNA
39
THE WHYrsquoS CUTTING PROTEIN AND DNA
The DNA in the nucleus of the cell is moulded folded and protected by proteins
40
THE WHYrsquoSALCOHOL Alcohol is less dense than water so it floats
on top Look for clumps of white stringy stuff where the water and alcohol layers meet
DNA is not soluble in alcohol - other cell parts are
By adding alcohol DNA precipitates out of the solution and collect at the interface of the alcohol and soap layer
The colder the alcohol the less soluble the DNA will be in it
41
COLD ALCOHOL
DNA dissolves in water but precipitates in alcohol
Cold alcohol is used to separate DNA out of water-based solutions
This allows the DNA to be purified for subsequent genetic testing
Adding alcohol to a solution containing DNA is a simple way to obtain the pure DNA required and colder temperatures slow down enzymes that can break down DNA giving better extraction results
42
WHY IS DNA EXTRACTION IMPORTANT DNA extraction is an important molecular
biology procedure
By definition extraction is taking DNA out of any type of cell for the purpose of analysis
See Handouts for more information on the use of DNA extracted (extension only)
43
SUMMARIZEDNA EXTRACTING FROM KIWI FRUIT
44
BREAK DOWN CELL WALLS
Cells walls have to be destroyed to reach the DNA within cells
Extraction procedures to obtain pure DNA have to get rid of all the molecular and chemical components of the tissue from which the DNA is being extracted
First steps involve lysing or destroying the cell walls This can be done with a variety of chemical agents that are caustic to cell membranes but do not harm the DNA Cells can also be sonicated homogenized or ground up to destroy membranes
45
REMOVING Removing Lipids
Once the cell walls have been destroyed a detergent is added to get rid of the fats and oils that make up the cell membranes Detergents cause the fats and oils to dissolve into the solution
Remove ProteinsProteins and enzymes can be digested by
adding a protease to the solution Proteases break down proteins into small peptides and amino acids
46
PRECIPITATING AND PURIFYING Precipitate DNA
Adding cold alcohol to a solution will cause DNA to precipitate and aggregate The DNA can be collected by centrifuging the sample and pouring off the liquid layer The DNA should exist as a small pellet in the bottom of the centrifuge tube
Purify DNAThe DNA can be washed by re-suspending it
in a cold alcohol solution and re-centrifuging it several times to obtain a very pure DNA sample Typical alcohols used to precipitate DNA include ethanol and Isopropanol This process leaves a very pure sample for stringent DNA testing
47
PART B OBSERVING EXTRACTED DNA UNDER THE MICROSCOPE
48
PART B A CLOSER LOOK
8 Use a dropping pipette to carefully remove some of the threadlike substance from the top of your preparation
9 Place one or two drops onto the middle of a
microscope slide
10 Add two drops of methylene blue Wait 3 or 4 minutes to allow the methylene blue to be absorbed by the DNA
11 Carefully place a cover slip on the slide Gently press
a folded piece of paper towelling over the top of the prepared slide to soak up any excess liquid
12 Observe the DNA under low power then high power
49
DISCUSSIONANSWER IN GROUP RELATED WORKED 1 Write a detailed description of the material
floating at the top of the test tube after the alcohol was added
2 Describe the DNA as it appears under the
microscope under high power 3 Prepare a diagram of your DNA specimen 4 What was the reason for using methylene
blue
50
HOMEWORKCONSTRUCT A FLOW CHART DRAWING MIND MAP OF THE PROCESS YOU USED EXTRACTING THE DNA FROM KIWIrsquoS
Next to each step explain how the method you used was important (Hint What substances make up the membranes of cells and cell organelles
How do detergents work
What is the active ingredient in meat tenderiser
51
TIPS STEPS FOR FLOW CHART In order to release the DNA from the
nuclei of the pea cells you first separated the cells from one another
Then the cell and nuclear membranes needed to be ruptured to release the cell contents and the contents of the nucleus
Once removed from the nuclear membrane the DNA had to be untangled into the visible threadlike structures you ended up with
52
HOMEWORK EXTENSION GROUP QUESTIONS Where can DNA be found in the cell Discuss the action of the soap (detergent) on the cell
What is the purpose of the soap in this activity What was the purpose of the Sodium Chloride Include a
discussion of polarity and charged particles Why was the cold ethanol added to the soap and salt
mixture Describe the appearance of your final product Draw a diagram of DNA containing 5 sets of nucleotide
bases labelling the hydrogen bonds between the bases References and Resources Adapted from Berry Full
of DNA by Diane Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
53
HTTPWWWGENOMEBCCAEDUCATIONTEACHERSCLASSROOM-ACTIVITIESDNA-EXTRACTION
54
EXTRACHROMOSOMES EXIST IN PAIRS Because our chromosomes exist in pairs (and consequently we
have 2 alleles of each gene) we are a diploid species This is why our somatic cells are represented as 2n Our gametes (sperm and ova) on the other hand are haploid and are represented as n Other species may have different ploidy for example
triploid (3n) seedless watermelons tetaploid (4n) salmonidae fish pentaploid (5n) Kenai birch hexaploid (6n) some types of wheat kiwi fruit octaploid (8n) acipenser (a genus
of sturgeon fish strawberies) decaploid (10n) some strawberries dodecaploid (12n) some types of amphibians eg Xenopus
ruwenzoriensis
wwwcyberusca
55
YOUR EXTRACTED DNA UNDER THE MICROSCOPE LOOKED
LIKE
Collaboration-Brainstorming on what to look at under the
- DNA extraction- DNA is too small for even a microscope to see and after the extraction it just appears like a blob True or false
Could you any DNA strand
wwwemployeescsbsjuedu
56
HANDOUT ON DNA EXTRACTIONBACKGROUND EVERY DAY LIFE
EXTRA READING MATERIAL(EXTENSION )
DISCOVERING DNA DNA ON THE INSIDE USE OF DNA EXTRACTION INN EVERYDAY LIFE WHY IS DNA TESTING GOOD MOLECULAR BIOLOGY PCR-based diagnostics of genomic DNA
57
REFERENCES httpwwwsquidoocomhow-to-get-dna-from-a-kiwi-fruit Why Is DNA Testing Good | eHowcom
httpwwwehowcomabout_6684410_dna-testing-good_htmlixzz1MkbcIJs5 Why Is DNA Extraction Important | eHowcom
httpwwwehowcomlist_5839095_dna-extraction-important_htmlixzz1MkZDrqyY
Why Is Sodium Used in DNA Extraction | eHowcom httpwwwehowcomabout_6504902_sodium-used-dna-extraction_htmlixzz1MkaGWg57
Why Is Cold Alcohol Used in DNA Tests | eHowcom httpwwwehowcomabout_6399349_cold-alcohol-used-dna-tests_htmlixzz1MkbEmEdu
httplearngeneticsutaheducontentlabsextractionhowto www kamibudaksainsblogspotcom
httpwwwchemwatch goldcom References and Resources Adapted from Berry Full of DNA by Diane
Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
58
REFERENCES
Uses of DNA Extraction | eHowcom httpwwwehowcomabout_5344428_uses-dna-extractionhtmlixzz1MkWQB2TZ
Science 10 A Contextual Approach Heineman Queensland Science Projects Regan Spence Maggie Spenceley
httpenwikipediaorgwikiMolecular_biology httpwwwclinchemorgcgicontent
extract44102201 http
employeescsbsjueduhjakubowskiclassesch331dnaoldnastructurehtml
httpwwwgenomebccaeducationteachersclassroom-activitiesdna-extraction
9
DNA extraction refers to the process that scientists use to break down a piece of evidence such as a piece of hair or a drop of blood in order to determine the individuals DNA which is unique to each person Understand the process of DNA extraction with information from a biology teacher in this free video on science
Expert Janice CrenettiContact WeAreHDTVcomBio Janice Creneti has a Bachelor of Science in secondary science education and a Bachelor of Art in biology from Boston UniversityFilmmaker Christopher Rokosz
Category Entertainment Tags science dna dna testing dna structure dna model dna replication dna molecule
10
REVIEWING OF INFORMATION ONLABORATORY SAFETY RULES AND RISK ASSESSMENTS
Do not enter the laboratory unless you are with a teacher Never touch equipment in the laboratory unless you are told to use it Donrsquot eat or drink in the laboratory Always walkmdashnever run Wear protective clothingmdash a laboratory coat or apron and when
appropriate safety glasses Never taste anything Tie up long hair Always point test tubes away from people Check with your teacher on how to dispose of waste liquids and solids
Broken glass should be cleaned up using gloves a brush and dustpan and placed in a special bin
If you spill something on your skin or clothes wash it immediately with lots of water Tell your teacher
Report all accidents and breakages to your teacher After heating equipment let it cool on a heatproof mat before picking
it up This will avoid burns Clean all equipment after use and put it back where you got it from
Clean and dry your work bench
11
RISK ASSESSMENT ON EXPERIMENT AND INVESTIGATION ndash KIWI FRUIT
Eyewear and protective clothingNo eat or drinkingCover the MSDSrsquo e on Isopropanol and Methylene blue ndash hard copy handouts (health and safety regulation)
Any broken glassware must be reported
Care must be taken when using and washing the blender ndash sharp blades
12
ISOPROPANOL ndash MSDS HAND OUTS CHEMWATCH ISSUE DATE 15052010 ISOPROPANOL - FLAMMABLE Dangerous Good 3 SAFETY-DATA(tm) Ratings (Provided here for your convenience)
----------------------------------------------------------------------------------------------------------- Health Rating 2 - Moderate Flammability Rating 3 - Severe (Flammable) Reactivity Rating 2 - Moderate Contact Rating 3 - Severe Lab Protective Equip GOGGLES amp SHIELD LAB COAT amp APRON VENT HOOD PROPER GLOVES CLASS B EXTINGUISHER Storage Colour Code Red (Flammable)
Potential Health Effects ----------------------------------
Inhalation Inhalation of vapours irritates the respiratory tract Exposure to high concentrations has a narcotic effect producing symptoms of dizziness drowsiness headache staggering unconsciousness and possibly death Ingestion Can cause drowsiness unconsciousness and death Gastrointestinal pain cramps nausea vomiting and diarrhoea may also result The single lethal dose for a human adult = about 250 mls (8 ounces) Skin Contact May cause irritation with redness and pain May be absorbed through the skin with possible systemic effects Eye Contact Vapours cause eye irritation Splashes cause severe irritation possible corneal burns and eye damage Chronic Exposure Chronic exposure may cause skin effects Aggravation of Pre-existing Conditions Persons with pre-existing skin disorders or impaired liver kidney or pulmonary function may be more susceptible to the effects of this agent
13
METHYLENE BLUE ndash MSDS HAND OUTS CHEMWATCH ISSUE DATE 15052010 METHYLENE BLUE SAFETY-DATA(tm) Ratings (Provided here for your convenience)
----------------------------------------------------------------------------------------------------------- Health Rating 2 - Moderate Flammability Rating 1 - Slight Reactivity Rating 1 - Slight Contact Rating 1 - Slight Lab Protective Equip GOGGLES LAB COAT VENT HOOD PROPER GLOVES Storage Colour Code Green (General Storage) -----------------------------------------------------------------------------------------------------------
Potential Health Effects ----------------------------------
This material is relatively nonhazardous in routine industrial situations
Inhalation No adverse health effects expected from inhalation May cause a short period of rapid or difficult breathing Ingestion A burning sensation of the mouth may be noted following ingestion of methylene blue May cause nausea vomiting diarrhea and gastritis Large doses may cause abdominal and chest pain headache profuse sweating mental confusion painful maturation and methemoglobinemia Skin Contact Not expected to be a health hazard from skin exposure Methylene blue may colour the skin a bluish colour May cause photosensitization Eye Contact No adverse effects expected May cause mechanical irritation Chronic Exposure No information found Aggravation of Pre-existing Conditions No information found
14
NEW WORDS TO WORD BANKADD THESE TO YOUR EXCEL ALPHA TABLE
Precipitate Spooling Denaturising Lysing Homogenized Aggregate Lipid Centrifuging PCR
WORDS MEANING
15
bullDO YOU THINK YOU HAVE VERY MUCH IN COMMON WITH A KIWI FRUIT
BELIEVE IT OR NOT A KIWIS GENETIC MATERIAL IS VERY SIMILAR TO YOUR OWN
SEE AND TOUCH THE GENETIC MATERIAL THAT YOULL EXTRACT FROM THE CELLS OF A KIWI FRUIT
Activity Overview
Extract DNA from kiwi fruit using simple household chemicals
16
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GEN
ETIC
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OW
TO
DNA EXTRACTION USING KIWI FRUIT
17
AIM
TO
EX
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OM
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IWI F
RU
IT
wwwexploratoriumedu
MATERIALS EQUIPMENT (per student group takes 30 minutes)
frac12 cup Kiwi fruit 200 mL water dishwashing detergent dropping pipette fine mesh kitchen strainer amp cheese cloth glass rod large beaker large test tube light microscope meat tenderiser methylene blue microscope lamp inoculation needle microscope slide and cover slip paper towelling small beaker of alcohol (Isopropanol) spatula test-tube rack mortar and pestle
18
PART A EXTRACTING THE DNA
METHOD
1 Peal and Place the kiwi and water in the beaker and mush it with a mortar
and pestle DNA SOURCE About 125 ml 12 cup Twice as much water as DNA source (250
ml 1 cup) Table salt large pinch 1g 14 teaspoon Stir until the mixture is of a thin soupy
consistency
TAKE NOTES ndash NO TEXTBOOKS ALLOWED WHEN PERFORMING EXPERIMENT ndashMUST RECALL PROCEDURES METHOD
19
WHAT MUST I DO NOW
STRAIN THE DNA MIXTURE
DISHWASHING DETERGENT
20
2 Pour the thin cell mixture through the kitchen strainer (cheesecloth) into another large beaker
3 Measure 12 ml of the soup into
a small beakerAdd 2 ml of liquid detergentSwirl to mix stir thoroughly using a
glass rodLet mixture sit in container with
hot tap water (60 - 65degC) for 5 - 10 minutes
21
WHAT MUST I DO NOWMEAT TENDERIZER
ENZYME POWDERALCOHOL SEPARATION
wwwforumsoverclockerscomau
22
4 Add a pinch of enzyme (meat tenderizer) to mixture
(Using pineapple juice or contact lens cleaning solution will do the same as tenderizer)
Gently stir with toothpick skewer for 5 minutes continue stirring not too vigorously Be careful If you stir too hard youll break up the DNA making it harder to see
5 Quarter-fill a large test tube with the mixture
23
SLOWLY pour the same amount ice - cold (70 - 95 isopropyl or ethyl alcohol) into test tube pour alcohol down the side of the test tube
Tilting the test tube will make this easier to do
It forms a layer on top of the cell mixture
DO NOT MIX THE TWO LAYERS TOGETHER
Amount of alcohol and mixture should be the same
24
SEPARATION USING ALCOHOL
25
7 Observe the mixture for a few minutes You will see a white threadlike substance rise from the mixture to rest above in the alcohol layer
This is the DNA that you have extracted from the cells of the kiwi fruit
8 You can get more DNA to precipitate from the solution using a DNA collecting tool (glass or paper clip hook or cut inoculation needle)
Gently lift the water solution up into the alcohol layer (this allows more DNA to get in contact with the alcohol and precipitate)
26
INFORMATION DNA precipitates as a white stringy
ldquosnottyrdquo film at the water - alcohol interface and eventually will rise into the alcohol layer from the mixture layer
Allow test tube to sit for several minutes The clearer the DNA is the fewer
impurities you have
If you have an acceptable amount of DNA it can be spooled by rotating your collecting tool and then transferred into a clean tube container
27
WOW ndash SPOOLING THE DNA If you are careful you may be able wind up the DNA
around a glass rod or a skewer Position the tip of the glass rod or skewer where you can see the threads of DNA Steadily twist the rod or skewer as if you were making candy floss Alternatively use a straw to pull it out by suction ndash be careful not to get in you mouth
Donrsquot go too quickly
You should be able to pull the strands of DNA out of the mixture
ASK STUDENTS TO TAKE MOBILE PHONE PICTURES OF THEIR OWN EXTRACTED DNA AND COMPARE APPEARANCE WITH OTHERS
Photos of steps can be inserted in flow diagram(ICT)
28
KIWIDNA
29
If you want to save your DNA you can transfer it to a small container filled with alcohol
Leave tube container uncapped until the ethanol has evaporated
DNA can be stored in the fridge (dry or water buffer can be added)
You can now investigate the property
30
WHAT IS THAT STRINGY STUFFDNA IS A LONG STRINGY MOLECULE THE SALT THAT YOU ADDED IN STEP ONE HELPS IT STICK TOGETHER SO WHAT YOU SEE ARE CLUMPS OF TANGLED DNA MOLECULES
DNA NORMALLY STAYS DISSOLVED IN WATER BUT WHEN SALTY DNA COMES IN CONTACT WITH ALCOHOL IT BECOMES UNDISSOLVED THIS IS CALLED PRECIPITATION THE PHYSICAL FORCE OF THE DNA CLUMPING TOGETHER AS IT PRECIPITATES PULLS MORE STRANDS ALONG WITH IT AS IT RISES INTO THE ALCOHOL
31
THE WHYrsquoS Blending separated the pea cells In order to extract DNA from a cell the associated
membranes and proteins must first be removed (break apart the cells) and then physically separated (loosen the tough cell wall) from the DNA
To see the DNA we have to break open these two sacks We do this with detergent and salt( Sodium can be
involved in several of the steps ) Sodium is an element Its chemical symbol is Na for
Natrium the Latin word for sodium It is a positive ion and often associates with negative ions as part of useful compounds
Salt help precipitate protein and carbohydrates away from the DNA
Salt helps strip away the proteins associated with DNA + and ndash charge
32
THE WHYrsquoS Salt and Detergents are used to break down
cell walls and nuclear membranes to release the DNA
They work by chemically poking holes in the cell membranes or walls
Once holes are poked in the membranes the membranes can be further disrupted mechanically as with a blender
After that it is easier to get the contents of the cell out including the DNA
LETS HAVE A LOOK AT THE CELL
33
CELL TO DNA
httplearngeneticsutaheducontentlabsextractionhowtoHOW TO EXTRACT DNA FROM ANYTHING LIVING
34
THE WHYrsquoS WHY DETERGENT
Each cell is surrounded by a sack (the cell membrane)
DNA is found inside the second sack (nucleus) within each cell
To see the DNA we have to break it open A cell membrane has 2 layers of lipid(fat)
molecules with protein going through them When the lysis buffer (detergent) comes
close to the cell it captures the lipids and the proteins - breaks open the cell destroying the fatty membrane - DNA now released into the solution
35
A CELLS MEMBRANES HAVE TWO LAYERS OF LIPID (FAT) MOLECULES WITH PROTEINS GOING THROUGH THEM
36
Why detergent How does detergent work
Think about why you use soap to wash dishes or your hands To remove grease and dirt right
Soap molecules and grease molecules are made of two parts
1(Blue) Heads which like water 2(Green) Tails which hate water
37
AFTER ADDING THE DETERGENT WHAT DO YOU HAVE IN YOUR SOUP
38
THE WHYrsquoS ENZYME (MEAT TENDERIZER)
The DNA in the nucleus of the cell is molded folded and protected by proteins The tenderizer cut the proteins away from the DNA
In this experiment meat tenderizer acts as an enzyme to cut proteins just like a pair of scissors
The meat tenderizer cuts the proteins away from the DNA
39
THE WHYrsquoS CUTTING PROTEIN AND DNA
The DNA in the nucleus of the cell is moulded folded and protected by proteins
40
THE WHYrsquoSALCOHOL Alcohol is less dense than water so it floats
on top Look for clumps of white stringy stuff where the water and alcohol layers meet
DNA is not soluble in alcohol - other cell parts are
By adding alcohol DNA precipitates out of the solution and collect at the interface of the alcohol and soap layer
The colder the alcohol the less soluble the DNA will be in it
41
COLD ALCOHOL
DNA dissolves in water but precipitates in alcohol
Cold alcohol is used to separate DNA out of water-based solutions
This allows the DNA to be purified for subsequent genetic testing
Adding alcohol to a solution containing DNA is a simple way to obtain the pure DNA required and colder temperatures slow down enzymes that can break down DNA giving better extraction results
42
WHY IS DNA EXTRACTION IMPORTANT DNA extraction is an important molecular
biology procedure
By definition extraction is taking DNA out of any type of cell for the purpose of analysis
See Handouts for more information on the use of DNA extracted (extension only)
43
SUMMARIZEDNA EXTRACTING FROM KIWI FRUIT
44
BREAK DOWN CELL WALLS
Cells walls have to be destroyed to reach the DNA within cells
Extraction procedures to obtain pure DNA have to get rid of all the molecular and chemical components of the tissue from which the DNA is being extracted
First steps involve lysing or destroying the cell walls This can be done with a variety of chemical agents that are caustic to cell membranes but do not harm the DNA Cells can also be sonicated homogenized or ground up to destroy membranes
45
REMOVING Removing Lipids
Once the cell walls have been destroyed a detergent is added to get rid of the fats and oils that make up the cell membranes Detergents cause the fats and oils to dissolve into the solution
Remove ProteinsProteins and enzymes can be digested by
adding a protease to the solution Proteases break down proteins into small peptides and amino acids
46
PRECIPITATING AND PURIFYING Precipitate DNA
Adding cold alcohol to a solution will cause DNA to precipitate and aggregate The DNA can be collected by centrifuging the sample and pouring off the liquid layer The DNA should exist as a small pellet in the bottom of the centrifuge tube
Purify DNAThe DNA can be washed by re-suspending it
in a cold alcohol solution and re-centrifuging it several times to obtain a very pure DNA sample Typical alcohols used to precipitate DNA include ethanol and Isopropanol This process leaves a very pure sample for stringent DNA testing
47
PART B OBSERVING EXTRACTED DNA UNDER THE MICROSCOPE
48
PART B A CLOSER LOOK
8 Use a dropping pipette to carefully remove some of the threadlike substance from the top of your preparation
9 Place one or two drops onto the middle of a
microscope slide
10 Add two drops of methylene blue Wait 3 or 4 minutes to allow the methylene blue to be absorbed by the DNA
11 Carefully place a cover slip on the slide Gently press
a folded piece of paper towelling over the top of the prepared slide to soak up any excess liquid
12 Observe the DNA under low power then high power
49
DISCUSSIONANSWER IN GROUP RELATED WORKED 1 Write a detailed description of the material
floating at the top of the test tube after the alcohol was added
2 Describe the DNA as it appears under the
microscope under high power 3 Prepare a diagram of your DNA specimen 4 What was the reason for using methylene
blue
50
HOMEWORKCONSTRUCT A FLOW CHART DRAWING MIND MAP OF THE PROCESS YOU USED EXTRACTING THE DNA FROM KIWIrsquoS
Next to each step explain how the method you used was important (Hint What substances make up the membranes of cells and cell organelles
How do detergents work
What is the active ingredient in meat tenderiser
51
TIPS STEPS FOR FLOW CHART In order to release the DNA from the
nuclei of the pea cells you first separated the cells from one another
Then the cell and nuclear membranes needed to be ruptured to release the cell contents and the contents of the nucleus
Once removed from the nuclear membrane the DNA had to be untangled into the visible threadlike structures you ended up with
52
HOMEWORK EXTENSION GROUP QUESTIONS Where can DNA be found in the cell Discuss the action of the soap (detergent) on the cell
What is the purpose of the soap in this activity What was the purpose of the Sodium Chloride Include a
discussion of polarity and charged particles Why was the cold ethanol added to the soap and salt
mixture Describe the appearance of your final product Draw a diagram of DNA containing 5 sets of nucleotide
bases labelling the hydrogen bonds between the bases References and Resources Adapted from Berry Full
of DNA by Diane Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
53
HTTPWWWGENOMEBCCAEDUCATIONTEACHERSCLASSROOM-ACTIVITIESDNA-EXTRACTION
54
EXTRACHROMOSOMES EXIST IN PAIRS Because our chromosomes exist in pairs (and consequently we
have 2 alleles of each gene) we are a diploid species This is why our somatic cells are represented as 2n Our gametes (sperm and ova) on the other hand are haploid and are represented as n Other species may have different ploidy for example
triploid (3n) seedless watermelons tetaploid (4n) salmonidae fish pentaploid (5n) Kenai birch hexaploid (6n) some types of wheat kiwi fruit octaploid (8n) acipenser (a genus
of sturgeon fish strawberies) decaploid (10n) some strawberries dodecaploid (12n) some types of amphibians eg Xenopus
ruwenzoriensis
wwwcyberusca
55
YOUR EXTRACTED DNA UNDER THE MICROSCOPE LOOKED
LIKE
Collaboration-Brainstorming on what to look at under the
- DNA extraction- DNA is too small for even a microscope to see and after the extraction it just appears like a blob True or false
Could you any DNA strand
wwwemployeescsbsjuedu
56
HANDOUT ON DNA EXTRACTIONBACKGROUND EVERY DAY LIFE
EXTRA READING MATERIAL(EXTENSION )
DISCOVERING DNA DNA ON THE INSIDE USE OF DNA EXTRACTION INN EVERYDAY LIFE WHY IS DNA TESTING GOOD MOLECULAR BIOLOGY PCR-based diagnostics of genomic DNA
57
REFERENCES httpwwwsquidoocomhow-to-get-dna-from-a-kiwi-fruit Why Is DNA Testing Good | eHowcom
httpwwwehowcomabout_6684410_dna-testing-good_htmlixzz1MkbcIJs5 Why Is DNA Extraction Important | eHowcom
httpwwwehowcomlist_5839095_dna-extraction-important_htmlixzz1MkZDrqyY
Why Is Sodium Used in DNA Extraction | eHowcom httpwwwehowcomabout_6504902_sodium-used-dna-extraction_htmlixzz1MkaGWg57
Why Is Cold Alcohol Used in DNA Tests | eHowcom httpwwwehowcomabout_6399349_cold-alcohol-used-dna-tests_htmlixzz1MkbEmEdu
httplearngeneticsutaheducontentlabsextractionhowto www kamibudaksainsblogspotcom
httpwwwchemwatch goldcom References and Resources Adapted from Berry Full of DNA by Diane
Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
58
REFERENCES
Uses of DNA Extraction | eHowcom httpwwwehowcomabout_5344428_uses-dna-extractionhtmlixzz1MkWQB2TZ
Science 10 A Contextual Approach Heineman Queensland Science Projects Regan Spence Maggie Spenceley
httpenwikipediaorgwikiMolecular_biology httpwwwclinchemorgcgicontent
extract44102201 http
employeescsbsjueduhjakubowskiclassesch331dnaoldnastructurehtml
httpwwwgenomebccaeducationteachersclassroom-activitiesdna-extraction
10
REVIEWING OF INFORMATION ONLABORATORY SAFETY RULES AND RISK ASSESSMENTS
Do not enter the laboratory unless you are with a teacher Never touch equipment in the laboratory unless you are told to use it Donrsquot eat or drink in the laboratory Always walkmdashnever run Wear protective clothingmdash a laboratory coat or apron and when
appropriate safety glasses Never taste anything Tie up long hair Always point test tubes away from people Check with your teacher on how to dispose of waste liquids and solids
Broken glass should be cleaned up using gloves a brush and dustpan and placed in a special bin
If you spill something on your skin or clothes wash it immediately with lots of water Tell your teacher
Report all accidents and breakages to your teacher After heating equipment let it cool on a heatproof mat before picking
it up This will avoid burns Clean all equipment after use and put it back where you got it from
Clean and dry your work bench
11
RISK ASSESSMENT ON EXPERIMENT AND INVESTIGATION ndash KIWI FRUIT
Eyewear and protective clothingNo eat or drinkingCover the MSDSrsquo e on Isopropanol and Methylene blue ndash hard copy handouts (health and safety regulation)
Any broken glassware must be reported
Care must be taken when using and washing the blender ndash sharp blades
12
ISOPROPANOL ndash MSDS HAND OUTS CHEMWATCH ISSUE DATE 15052010 ISOPROPANOL - FLAMMABLE Dangerous Good 3 SAFETY-DATA(tm) Ratings (Provided here for your convenience)
----------------------------------------------------------------------------------------------------------- Health Rating 2 - Moderate Flammability Rating 3 - Severe (Flammable) Reactivity Rating 2 - Moderate Contact Rating 3 - Severe Lab Protective Equip GOGGLES amp SHIELD LAB COAT amp APRON VENT HOOD PROPER GLOVES CLASS B EXTINGUISHER Storage Colour Code Red (Flammable)
Potential Health Effects ----------------------------------
Inhalation Inhalation of vapours irritates the respiratory tract Exposure to high concentrations has a narcotic effect producing symptoms of dizziness drowsiness headache staggering unconsciousness and possibly death Ingestion Can cause drowsiness unconsciousness and death Gastrointestinal pain cramps nausea vomiting and diarrhoea may also result The single lethal dose for a human adult = about 250 mls (8 ounces) Skin Contact May cause irritation with redness and pain May be absorbed through the skin with possible systemic effects Eye Contact Vapours cause eye irritation Splashes cause severe irritation possible corneal burns and eye damage Chronic Exposure Chronic exposure may cause skin effects Aggravation of Pre-existing Conditions Persons with pre-existing skin disorders or impaired liver kidney or pulmonary function may be more susceptible to the effects of this agent
13
METHYLENE BLUE ndash MSDS HAND OUTS CHEMWATCH ISSUE DATE 15052010 METHYLENE BLUE SAFETY-DATA(tm) Ratings (Provided here for your convenience)
----------------------------------------------------------------------------------------------------------- Health Rating 2 - Moderate Flammability Rating 1 - Slight Reactivity Rating 1 - Slight Contact Rating 1 - Slight Lab Protective Equip GOGGLES LAB COAT VENT HOOD PROPER GLOVES Storage Colour Code Green (General Storage) -----------------------------------------------------------------------------------------------------------
Potential Health Effects ----------------------------------
This material is relatively nonhazardous in routine industrial situations
Inhalation No adverse health effects expected from inhalation May cause a short period of rapid or difficult breathing Ingestion A burning sensation of the mouth may be noted following ingestion of methylene blue May cause nausea vomiting diarrhea and gastritis Large doses may cause abdominal and chest pain headache profuse sweating mental confusion painful maturation and methemoglobinemia Skin Contact Not expected to be a health hazard from skin exposure Methylene blue may colour the skin a bluish colour May cause photosensitization Eye Contact No adverse effects expected May cause mechanical irritation Chronic Exposure No information found Aggravation of Pre-existing Conditions No information found
14
NEW WORDS TO WORD BANKADD THESE TO YOUR EXCEL ALPHA TABLE
Precipitate Spooling Denaturising Lysing Homogenized Aggregate Lipid Centrifuging PCR
WORDS MEANING
15
bullDO YOU THINK YOU HAVE VERY MUCH IN COMMON WITH A KIWI FRUIT
BELIEVE IT OR NOT A KIWIS GENETIC MATERIAL IS VERY SIMILAR TO YOUR OWN
SEE AND TOUCH THE GENETIC MATERIAL THAT YOULL EXTRACT FROM THE CELLS OF A KIWI FRUIT
Activity Overview
Extract DNA from kiwi fruit using simple household chemicals
16
HT
TP
LEA
RN
GEN
ETIC
SU
TA
HE
DU
CO
NTE
NTL
AB
SE
XTR
AC
TIO
NH
OW
TO
DNA EXTRACTION USING KIWI FRUIT
17
AIM
TO
EX
TR
AC
T D
NA
FR
OM
TH
E C
ELLS
OF K
IWI F
RU
IT
wwwexploratoriumedu
MATERIALS EQUIPMENT (per student group takes 30 minutes)
frac12 cup Kiwi fruit 200 mL water dishwashing detergent dropping pipette fine mesh kitchen strainer amp cheese cloth glass rod large beaker large test tube light microscope meat tenderiser methylene blue microscope lamp inoculation needle microscope slide and cover slip paper towelling small beaker of alcohol (Isopropanol) spatula test-tube rack mortar and pestle
18
PART A EXTRACTING THE DNA
METHOD
1 Peal and Place the kiwi and water in the beaker and mush it with a mortar
and pestle DNA SOURCE About 125 ml 12 cup Twice as much water as DNA source (250
ml 1 cup) Table salt large pinch 1g 14 teaspoon Stir until the mixture is of a thin soupy
consistency
TAKE NOTES ndash NO TEXTBOOKS ALLOWED WHEN PERFORMING EXPERIMENT ndashMUST RECALL PROCEDURES METHOD
19
WHAT MUST I DO NOW
STRAIN THE DNA MIXTURE
DISHWASHING DETERGENT
20
2 Pour the thin cell mixture through the kitchen strainer (cheesecloth) into another large beaker
3 Measure 12 ml of the soup into
a small beakerAdd 2 ml of liquid detergentSwirl to mix stir thoroughly using a
glass rodLet mixture sit in container with
hot tap water (60 - 65degC) for 5 - 10 minutes
21
WHAT MUST I DO NOWMEAT TENDERIZER
ENZYME POWDERALCOHOL SEPARATION
wwwforumsoverclockerscomau
22
4 Add a pinch of enzyme (meat tenderizer) to mixture
(Using pineapple juice or contact lens cleaning solution will do the same as tenderizer)
Gently stir with toothpick skewer for 5 minutes continue stirring not too vigorously Be careful If you stir too hard youll break up the DNA making it harder to see
5 Quarter-fill a large test tube with the mixture
23
SLOWLY pour the same amount ice - cold (70 - 95 isopropyl or ethyl alcohol) into test tube pour alcohol down the side of the test tube
Tilting the test tube will make this easier to do
It forms a layer on top of the cell mixture
DO NOT MIX THE TWO LAYERS TOGETHER
Amount of alcohol and mixture should be the same
24
SEPARATION USING ALCOHOL
25
7 Observe the mixture for a few minutes You will see a white threadlike substance rise from the mixture to rest above in the alcohol layer
This is the DNA that you have extracted from the cells of the kiwi fruit
8 You can get more DNA to precipitate from the solution using a DNA collecting tool (glass or paper clip hook or cut inoculation needle)
Gently lift the water solution up into the alcohol layer (this allows more DNA to get in contact with the alcohol and precipitate)
26
INFORMATION DNA precipitates as a white stringy
ldquosnottyrdquo film at the water - alcohol interface and eventually will rise into the alcohol layer from the mixture layer
Allow test tube to sit for several minutes The clearer the DNA is the fewer
impurities you have
If you have an acceptable amount of DNA it can be spooled by rotating your collecting tool and then transferred into a clean tube container
27
WOW ndash SPOOLING THE DNA If you are careful you may be able wind up the DNA
around a glass rod or a skewer Position the tip of the glass rod or skewer where you can see the threads of DNA Steadily twist the rod or skewer as if you were making candy floss Alternatively use a straw to pull it out by suction ndash be careful not to get in you mouth
Donrsquot go too quickly
You should be able to pull the strands of DNA out of the mixture
ASK STUDENTS TO TAKE MOBILE PHONE PICTURES OF THEIR OWN EXTRACTED DNA AND COMPARE APPEARANCE WITH OTHERS
Photos of steps can be inserted in flow diagram(ICT)
28
KIWIDNA
29
If you want to save your DNA you can transfer it to a small container filled with alcohol
Leave tube container uncapped until the ethanol has evaporated
DNA can be stored in the fridge (dry or water buffer can be added)
You can now investigate the property
30
WHAT IS THAT STRINGY STUFFDNA IS A LONG STRINGY MOLECULE THE SALT THAT YOU ADDED IN STEP ONE HELPS IT STICK TOGETHER SO WHAT YOU SEE ARE CLUMPS OF TANGLED DNA MOLECULES
DNA NORMALLY STAYS DISSOLVED IN WATER BUT WHEN SALTY DNA COMES IN CONTACT WITH ALCOHOL IT BECOMES UNDISSOLVED THIS IS CALLED PRECIPITATION THE PHYSICAL FORCE OF THE DNA CLUMPING TOGETHER AS IT PRECIPITATES PULLS MORE STRANDS ALONG WITH IT AS IT RISES INTO THE ALCOHOL
31
THE WHYrsquoS Blending separated the pea cells In order to extract DNA from a cell the associated
membranes and proteins must first be removed (break apart the cells) and then physically separated (loosen the tough cell wall) from the DNA
To see the DNA we have to break open these two sacks We do this with detergent and salt( Sodium can be
involved in several of the steps ) Sodium is an element Its chemical symbol is Na for
Natrium the Latin word for sodium It is a positive ion and often associates with negative ions as part of useful compounds
Salt help precipitate protein and carbohydrates away from the DNA
Salt helps strip away the proteins associated with DNA + and ndash charge
32
THE WHYrsquoS Salt and Detergents are used to break down
cell walls and nuclear membranes to release the DNA
They work by chemically poking holes in the cell membranes or walls
Once holes are poked in the membranes the membranes can be further disrupted mechanically as with a blender
After that it is easier to get the contents of the cell out including the DNA
LETS HAVE A LOOK AT THE CELL
33
CELL TO DNA
httplearngeneticsutaheducontentlabsextractionhowtoHOW TO EXTRACT DNA FROM ANYTHING LIVING
34
THE WHYrsquoS WHY DETERGENT
Each cell is surrounded by a sack (the cell membrane)
DNA is found inside the second sack (nucleus) within each cell
To see the DNA we have to break it open A cell membrane has 2 layers of lipid(fat)
molecules with protein going through them When the lysis buffer (detergent) comes
close to the cell it captures the lipids and the proteins - breaks open the cell destroying the fatty membrane - DNA now released into the solution
35
A CELLS MEMBRANES HAVE TWO LAYERS OF LIPID (FAT) MOLECULES WITH PROTEINS GOING THROUGH THEM
36
Why detergent How does detergent work
Think about why you use soap to wash dishes or your hands To remove grease and dirt right
Soap molecules and grease molecules are made of two parts
1(Blue) Heads which like water 2(Green) Tails which hate water
37
AFTER ADDING THE DETERGENT WHAT DO YOU HAVE IN YOUR SOUP
38
THE WHYrsquoS ENZYME (MEAT TENDERIZER)
The DNA in the nucleus of the cell is molded folded and protected by proteins The tenderizer cut the proteins away from the DNA
In this experiment meat tenderizer acts as an enzyme to cut proteins just like a pair of scissors
The meat tenderizer cuts the proteins away from the DNA
39
THE WHYrsquoS CUTTING PROTEIN AND DNA
The DNA in the nucleus of the cell is moulded folded and protected by proteins
40
THE WHYrsquoSALCOHOL Alcohol is less dense than water so it floats
on top Look for clumps of white stringy stuff where the water and alcohol layers meet
DNA is not soluble in alcohol - other cell parts are
By adding alcohol DNA precipitates out of the solution and collect at the interface of the alcohol and soap layer
The colder the alcohol the less soluble the DNA will be in it
41
COLD ALCOHOL
DNA dissolves in water but precipitates in alcohol
Cold alcohol is used to separate DNA out of water-based solutions
This allows the DNA to be purified for subsequent genetic testing
Adding alcohol to a solution containing DNA is a simple way to obtain the pure DNA required and colder temperatures slow down enzymes that can break down DNA giving better extraction results
42
WHY IS DNA EXTRACTION IMPORTANT DNA extraction is an important molecular
biology procedure
By definition extraction is taking DNA out of any type of cell for the purpose of analysis
See Handouts for more information on the use of DNA extracted (extension only)
43
SUMMARIZEDNA EXTRACTING FROM KIWI FRUIT
44
BREAK DOWN CELL WALLS
Cells walls have to be destroyed to reach the DNA within cells
Extraction procedures to obtain pure DNA have to get rid of all the molecular and chemical components of the tissue from which the DNA is being extracted
First steps involve lysing or destroying the cell walls This can be done with a variety of chemical agents that are caustic to cell membranes but do not harm the DNA Cells can also be sonicated homogenized or ground up to destroy membranes
45
REMOVING Removing Lipids
Once the cell walls have been destroyed a detergent is added to get rid of the fats and oils that make up the cell membranes Detergents cause the fats and oils to dissolve into the solution
Remove ProteinsProteins and enzymes can be digested by
adding a protease to the solution Proteases break down proteins into small peptides and amino acids
46
PRECIPITATING AND PURIFYING Precipitate DNA
Adding cold alcohol to a solution will cause DNA to precipitate and aggregate The DNA can be collected by centrifuging the sample and pouring off the liquid layer The DNA should exist as a small pellet in the bottom of the centrifuge tube
Purify DNAThe DNA can be washed by re-suspending it
in a cold alcohol solution and re-centrifuging it several times to obtain a very pure DNA sample Typical alcohols used to precipitate DNA include ethanol and Isopropanol This process leaves a very pure sample for stringent DNA testing
47
PART B OBSERVING EXTRACTED DNA UNDER THE MICROSCOPE
48
PART B A CLOSER LOOK
8 Use a dropping pipette to carefully remove some of the threadlike substance from the top of your preparation
9 Place one or two drops onto the middle of a
microscope slide
10 Add two drops of methylene blue Wait 3 or 4 minutes to allow the methylene blue to be absorbed by the DNA
11 Carefully place a cover slip on the slide Gently press
a folded piece of paper towelling over the top of the prepared slide to soak up any excess liquid
12 Observe the DNA under low power then high power
49
DISCUSSIONANSWER IN GROUP RELATED WORKED 1 Write a detailed description of the material
floating at the top of the test tube after the alcohol was added
2 Describe the DNA as it appears under the
microscope under high power 3 Prepare a diagram of your DNA specimen 4 What was the reason for using methylene
blue
50
HOMEWORKCONSTRUCT A FLOW CHART DRAWING MIND MAP OF THE PROCESS YOU USED EXTRACTING THE DNA FROM KIWIrsquoS
Next to each step explain how the method you used was important (Hint What substances make up the membranes of cells and cell organelles
How do detergents work
What is the active ingredient in meat tenderiser
51
TIPS STEPS FOR FLOW CHART In order to release the DNA from the
nuclei of the pea cells you first separated the cells from one another
Then the cell and nuclear membranes needed to be ruptured to release the cell contents and the contents of the nucleus
Once removed from the nuclear membrane the DNA had to be untangled into the visible threadlike structures you ended up with
52
HOMEWORK EXTENSION GROUP QUESTIONS Where can DNA be found in the cell Discuss the action of the soap (detergent) on the cell
What is the purpose of the soap in this activity What was the purpose of the Sodium Chloride Include a
discussion of polarity and charged particles Why was the cold ethanol added to the soap and salt
mixture Describe the appearance of your final product Draw a diagram of DNA containing 5 sets of nucleotide
bases labelling the hydrogen bonds between the bases References and Resources Adapted from Berry Full
of DNA by Diane Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
53
HTTPWWWGENOMEBCCAEDUCATIONTEACHERSCLASSROOM-ACTIVITIESDNA-EXTRACTION
54
EXTRACHROMOSOMES EXIST IN PAIRS Because our chromosomes exist in pairs (and consequently we
have 2 alleles of each gene) we are a diploid species This is why our somatic cells are represented as 2n Our gametes (sperm and ova) on the other hand are haploid and are represented as n Other species may have different ploidy for example
triploid (3n) seedless watermelons tetaploid (4n) salmonidae fish pentaploid (5n) Kenai birch hexaploid (6n) some types of wheat kiwi fruit octaploid (8n) acipenser (a genus
of sturgeon fish strawberies) decaploid (10n) some strawberries dodecaploid (12n) some types of amphibians eg Xenopus
ruwenzoriensis
wwwcyberusca
55
YOUR EXTRACTED DNA UNDER THE MICROSCOPE LOOKED
LIKE
Collaboration-Brainstorming on what to look at under the
- DNA extraction- DNA is too small for even a microscope to see and after the extraction it just appears like a blob True or false
Could you any DNA strand
wwwemployeescsbsjuedu
56
HANDOUT ON DNA EXTRACTIONBACKGROUND EVERY DAY LIFE
EXTRA READING MATERIAL(EXTENSION )
DISCOVERING DNA DNA ON THE INSIDE USE OF DNA EXTRACTION INN EVERYDAY LIFE WHY IS DNA TESTING GOOD MOLECULAR BIOLOGY PCR-based diagnostics of genomic DNA
57
REFERENCES httpwwwsquidoocomhow-to-get-dna-from-a-kiwi-fruit Why Is DNA Testing Good | eHowcom
httpwwwehowcomabout_6684410_dna-testing-good_htmlixzz1MkbcIJs5 Why Is DNA Extraction Important | eHowcom
httpwwwehowcomlist_5839095_dna-extraction-important_htmlixzz1MkZDrqyY
Why Is Sodium Used in DNA Extraction | eHowcom httpwwwehowcomabout_6504902_sodium-used-dna-extraction_htmlixzz1MkaGWg57
Why Is Cold Alcohol Used in DNA Tests | eHowcom httpwwwehowcomabout_6399349_cold-alcohol-used-dna-tests_htmlixzz1MkbEmEdu
httplearngeneticsutaheducontentlabsextractionhowto www kamibudaksainsblogspotcom
httpwwwchemwatch goldcom References and Resources Adapted from Berry Full of DNA by Diane
Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
58
REFERENCES
Uses of DNA Extraction | eHowcom httpwwwehowcomabout_5344428_uses-dna-extractionhtmlixzz1MkWQB2TZ
Science 10 A Contextual Approach Heineman Queensland Science Projects Regan Spence Maggie Spenceley
httpenwikipediaorgwikiMolecular_biology httpwwwclinchemorgcgicontent
extract44102201 http
employeescsbsjueduhjakubowskiclassesch331dnaoldnastructurehtml
httpwwwgenomebccaeducationteachersclassroom-activitiesdna-extraction
11
RISK ASSESSMENT ON EXPERIMENT AND INVESTIGATION ndash KIWI FRUIT
Eyewear and protective clothingNo eat or drinkingCover the MSDSrsquo e on Isopropanol and Methylene blue ndash hard copy handouts (health and safety regulation)
Any broken glassware must be reported
Care must be taken when using and washing the blender ndash sharp blades
12
ISOPROPANOL ndash MSDS HAND OUTS CHEMWATCH ISSUE DATE 15052010 ISOPROPANOL - FLAMMABLE Dangerous Good 3 SAFETY-DATA(tm) Ratings (Provided here for your convenience)
----------------------------------------------------------------------------------------------------------- Health Rating 2 - Moderate Flammability Rating 3 - Severe (Flammable) Reactivity Rating 2 - Moderate Contact Rating 3 - Severe Lab Protective Equip GOGGLES amp SHIELD LAB COAT amp APRON VENT HOOD PROPER GLOVES CLASS B EXTINGUISHER Storage Colour Code Red (Flammable)
Potential Health Effects ----------------------------------
Inhalation Inhalation of vapours irritates the respiratory tract Exposure to high concentrations has a narcotic effect producing symptoms of dizziness drowsiness headache staggering unconsciousness and possibly death Ingestion Can cause drowsiness unconsciousness and death Gastrointestinal pain cramps nausea vomiting and diarrhoea may also result The single lethal dose for a human adult = about 250 mls (8 ounces) Skin Contact May cause irritation with redness and pain May be absorbed through the skin with possible systemic effects Eye Contact Vapours cause eye irritation Splashes cause severe irritation possible corneal burns and eye damage Chronic Exposure Chronic exposure may cause skin effects Aggravation of Pre-existing Conditions Persons with pre-existing skin disorders or impaired liver kidney or pulmonary function may be more susceptible to the effects of this agent
13
METHYLENE BLUE ndash MSDS HAND OUTS CHEMWATCH ISSUE DATE 15052010 METHYLENE BLUE SAFETY-DATA(tm) Ratings (Provided here for your convenience)
----------------------------------------------------------------------------------------------------------- Health Rating 2 - Moderate Flammability Rating 1 - Slight Reactivity Rating 1 - Slight Contact Rating 1 - Slight Lab Protective Equip GOGGLES LAB COAT VENT HOOD PROPER GLOVES Storage Colour Code Green (General Storage) -----------------------------------------------------------------------------------------------------------
Potential Health Effects ----------------------------------
This material is relatively nonhazardous in routine industrial situations
Inhalation No adverse health effects expected from inhalation May cause a short period of rapid or difficult breathing Ingestion A burning sensation of the mouth may be noted following ingestion of methylene blue May cause nausea vomiting diarrhea and gastritis Large doses may cause abdominal and chest pain headache profuse sweating mental confusion painful maturation and methemoglobinemia Skin Contact Not expected to be a health hazard from skin exposure Methylene blue may colour the skin a bluish colour May cause photosensitization Eye Contact No adverse effects expected May cause mechanical irritation Chronic Exposure No information found Aggravation of Pre-existing Conditions No information found
14
NEW WORDS TO WORD BANKADD THESE TO YOUR EXCEL ALPHA TABLE
Precipitate Spooling Denaturising Lysing Homogenized Aggregate Lipid Centrifuging PCR
WORDS MEANING
15
bullDO YOU THINK YOU HAVE VERY MUCH IN COMMON WITH A KIWI FRUIT
BELIEVE IT OR NOT A KIWIS GENETIC MATERIAL IS VERY SIMILAR TO YOUR OWN
SEE AND TOUCH THE GENETIC MATERIAL THAT YOULL EXTRACT FROM THE CELLS OF A KIWI FRUIT
Activity Overview
Extract DNA from kiwi fruit using simple household chemicals
16
HT
TP
LEA
RN
GEN
ETIC
SU
TA
HE
DU
CO
NTE
NTL
AB
SE
XTR
AC
TIO
NH
OW
TO
DNA EXTRACTION USING KIWI FRUIT
17
AIM
TO
EX
TR
AC
T D
NA
FR
OM
TH
E C
ELLS
OF K
IWI F
RU
IT
wwwexploratoriumedu
MATERIALS EQUIPMENT (per student group takes 30 minutes)
frac12 cup Kiwi fruit 200 mL water dishwashing detergent dropping pipette fine mesh kitchen strainer amp cheese cloth glass rod large beaker large test tube light microscope meat tenderiser methylene blue microscope lamp inoculation needle microscope slide and cover slip paper towelling small beaker of alcohol (Isopropanol) spatula test-tube rack mortar and pestle
18
PART A EXTRACTING THE DNA
METHOD
1 Peal and Place the kiwi and water in the beaker and mush it with a mortar
and pestle DNA SOURCE About 125 ml 12 cup Twice as much water as DNA source (250
ml 1 cup) Table salt large pinch 1g 14 teaspoon Stir until the mixture is of a thin soupy
consistency
TAKE NOTES ndash NO TEXTBOOKS ALLOWED WHEN PERFORMING EXPERIMENT ndashMUST RECALL PROCEDURES METHOD
19
WHAT MUST I DO NOW
STRAIN THE DNA MIXTURE
DISHWASHING DETERGENT
20
2 Pour the thin cell mixture through the kitchen strainer (cheesecloth) into another large beaker
3 Measure 12 ml of the soup into
a small beakerAdd 2 ml of liquid detergentSwirl to mix stir thoroughly using a
glass rodLet mixture sit in container with
hot tap water (60 - 65degC) for 5 - 10 minutes
21
WHAT MUST I DO NOWMEAT TENDERIZER
ENZYME POWDERALCOHOL SEPARATION
wwwforumsoverclockerscomau
22
4 Add a pinch of enzyme (meat tenderizer) to mixture
(Using pineapple juice or contact lens cleaning solution will do the same as tenderizer)
Gently stir with toothpick skewer for 5 minutes continue stirring not too vigorously Be careful If you stir too hard youll break up the DNA making it harder to see
5 Quarter-fill a large test tube with the mixture
23
SLOWLY pour the same amount ice - cold (70 - 95 isopropyl or ethyl alcohol) into test tube pour alcohol down the side of the test tube
Tilting the test tube will make this easier to do
It forms a layer on top of the cell mixture
DO NOT MIX THE TWO LAYERS TOGETHER
Amount of alcohol and mixture should be the same
24
SEPARATION USING ALCOHOL
25
7 Observe the mixture for a few minutes You will see a white threadlike substance rise from the mixture to rest above in the alcohol layer
This is the DNA that you have extracted from the cells of the kiwi fruit
8 You can get more DNA to precipitate from the solution using a DNA collecting tool (glass or paper clip hook or cut inoculation needle)
Gently lift the water solution up into the alcohol layer (this allows more DNA to get in contact with the alcohol and precipitate)
26
INFORMATION DNA precipitates as a white stringy
ldquosnottyrdquo film at the water - alcohol interface and eventually will rise into the alcohol layer from the mixture layer
Allow test tube to sit for several minutes The clearer the DNA is the fewer
impurities you have
If you have an acceptable amount of DNA it can be spooled by rotating your collecting tool and then transferred into a clean tube container
27
WOW ndash SPOOLING THE DNA If you are careful you may be able wind up the DNA
around a glass rod or a skewer Position the tip of the glass rod or skewer where you can see the threads of DNA Steadily twist the rod or skewer as if you were making candy floss Alternatively use a straw to pull it out by suction ndash be careful not to get in you mouth
Donrsquot go too quickly
You should be able to pull the strands of DNA out of the mixture
ASK STUDENTS TO TAKE MOBILE PHONE PICTURES OF THEIR OWN EXTRACTED DNA AND COMPARE APPEARANCE WITH OTHERS
Photos of steps can be inserted in flow diagram(ICT)
28
KIWIDNA
29
If you want to save your DNA you can transfer it to a small container filled with alcohol
Leave tube container uncapped until the ethanol has evaporated
DNA can be stored in the fridge (dry or water buffer can be added)
You can now investigate the property
30
WHAT IS THAT STRINGY STUFFDNA IS A LONG STRINGY MOLECULE THE SALT THAT YOU ADDED IN STEP ONE HELPS IT STICK TOGETHER SO WHAT YOU SEE ARE CLUMPS OF TANGLED DNA MOLECULES
DNA NORMALLY STAYS DISSOLVED IN WATER BUT WHEN SALTY DNA COMES IN CONTACT WITH ALCOHOL IT BECOMES UNDISSOLVED THIS IS CALLED PRECIPITATION THE PHYSICAL FORCE OF THE DNA CLUMPING TOGETHER AS IT PRECIPITATES PULLS MORE STRANDS ALONG WITH IT AS IT RISES INTO THE ALCOHOL
31
THE WHYrsquoS Blending separated the pea cells In order to extract DNA from a cell the associated
membranes and proteins must first be removed (break apart the cells) and then physically separated (loosen the tough cell wall) from the DNA
To see the DNA we have to break open these two sacks We do this with detergent and salt( Sodium can be
involved in several of the steps ) Sodium is an element Its chemical symbol is Na for
Natrium the Latin word for sodium It is a positive ion and often associates with negative ions as part of useful compounds
Salt help precipitate protein and carbohydrates away from the DNA
Salt helps strip away the proteins associated with DNA + and ndash charge
32
THE WHYrsquoS Salt and Detergents are used to break down
cell walls and nuclear membranes to release the DNA
They work by chemically poking holes in the cell membranes or walls
Once holes are poked in the membranes the membranes can be further disrupted mechanically as with a blender
After that it is easier to get the contents of the cell out including the DNA
LETS HAVE A LOOK AT THE CELL
33
CELL TO DNA
httplearngeneticsutaheducontentlabsextractionhowtoHOW TO EXTRACT DNA FROM ANYTHING LIVING
34
THE WHYrsquoS WHY DETERGENT
Each cell is surrounded by a sack (the cell membrane)
DNA is found inside the second sack (nucleus) within each cell
To see the DNA we have to break it open A cell membrane has 2 layers of lipid(fat)
molecules with protein going through them When the lysis buffer (detergent) comes
close to the cell it captures the lipids and the proteins - breaks open the cell destroying the fatty membrane - DNA now released into the solution
35
A CELLS MEMBRANES HAVE TWO LAYERS OF LIPID (FAT) MOLECULES WITH PROTEINS GOING THROUGH THEM
36
Why detergent How does detergent work
Think about why you use soap to wash dishes or your hands To remove grease and dirt right
Soap molecules and grease molecules are made of two parts
1(Blue) Heads which like water 2(Green) Tails which hate water
37
AFTER ADDING THE DETERGENT WHAT DO YOU HAVE IN YOUR SOUP
38
THE WHYrsquoS ENZYME (MEAT TENDERIZER)
The DNA in the nucleus of the cell is molded folded and protected by proteins The tenderizer cut the proteins away from the DNA
In this experiment meat tenderizer acts as an enzyme to cut proteins just like a pair of scissors
The meat tenderizer cuts the proteins away from the DNA
39
THE WHYrsquoS CUTTING PROTEIN AND DNA
The DNA in the nucleus of the cell is moulded folded and protected by proteins
40
THE WHYrsquoSALCOHOL Alcohol is less dense than water so it floats
on top Look for clumps of white stringy stuff where the water and alcohol layers meet
DNA is not soluble in alcohol - other cell parts are
By adding alcohol DNA precipitates out of the solution and collect at the interface of the alcohol and soap layer
The colder the alcohol the less soluble the DNA will be in it
41
COLD ALCOHOL
DNA dissolves in water but precipitates in alcohol
Cold alcohol is used to separate DNA out of water-based solutions
This allows the DNA to be purified for subsequent genetic testing
Adding alcohol to a solution containing DNA is a simple way to obtain the pure DNA required and colder temperatures slow down enzymes that can break down DNA giving better extraction results
42
WHY IS DNA EXTRACTION IMPORTANT DNA extraction is an important molecular
biology procedure
By definition extraction is taking DNA out of any type of cell for the purpose of analysis
See Handouts for more information on the use of DNA extracted (extension only)
43
SUMMARIZEDNA EXTRACTING FROM KIWI FRUIT
44
BREAK DOWN CELL WALLS
Cells walls have to be destroyed to reach the DNA within cells
Extraction procedures to obtain pure DNA have to get rid of all the molecular and chemical components of the tissue from which the DNA is being extracted
First steps involve lysing or destroying the cell walls This can be done with a variety of chemical agents that are caustic to cell membranes but do not harm the DNA Cells can also be sonicated homogenized or ground up to destroy membranes
45
REMOVING Removing Lipids
Once the cell walls have been destroyed a detergent is added to get rid of the fats and oils that make up the cell membranes Detergents cause the fats and oils to dissolve into the solution
Remove ProteinsProteins and enzymes can be digested by
adding a protease to the solution Proteases break down proteins into small peptides and amino acids
46
PRECIPITATING AND PURIFYING Precipitate DNA
Adding cold alcohol to a solution will cause DNA to precipitate and aggregate The DNA can be collected by centrifuging the sample and pouring off the liquid layer The DNA should exist as a small pellet in the bottom of the centrifuge tube
Purify DNAThe DNA can be washed by re-suspending it
in a cold alcohol solution and re-centrifuging it several times to obtain a very pure DNA sample Typical alcohols used to precipitate DNA include ethanol and Isopropanol This process leaves a very pure sample for stringent DNA testing
47
PART B OBSERVING EXTRACTED DNA UNDER THE MICROSCOPE
48
PART B A CLOSER LOOK
8 Use a dropping pipette to carefully remove some of the threadlike substance from the top of your preparation
9 Place one or two drops onto the middle of a
microscope slide
10 Add two drops of methylene blue Wait 3 or 4 minutes to allow the methylene blue to be absorbed by the DNA
11 Carefully place a cover slip on the slide Gently press
a folded piece of paper towelling over the top of the prepared slide to soak up any excess liquid
12 Observe the DNA under low power then high power
49
DISCUSSIONANSWER IN GROUP RELATED WORKED 1 Write a detailed description of the material
floating at the top of the test tube after the alcohol was added
2 Describe the DNA as it appears under the
microscope under high power 3 Prepare a diagram of your DNA specimen 4 What was the reason for using methylene
blue
50
HOMEWORKCONSTRUCT A FLOW CHART DRAWING MIND MAP OF THE PROCESS YOU USED EXTRACTING THE DNA FROM KIWIrsquoS
Next to each step explain how the method you used was important (Hint What substances make up the membranes of cells and cell organelles
How do detergents work
What is the active ingredient in meat tenderiser
51
TIPS STEPS FOR FLOW CHART In order to release the DNA from the
nuclei of the pea cells you first separated the cells from one another
Then the cell and nuclear membranes needed to be ruptured to release the cell contents and the contents of the nucleus
Once removed from the nuclear membrane the DNA had to be untangled into the visible threadlike structures you ended up with
52
HOMEWORK EXTENSION GROUP QUESTIONS Where can DNA be found in the cell Discuss the action of the soap (detergent) on the cell
What is the purpose of the soap in this activity What was the purpose of the Sodium Chloride Include a
discussion of polarity and charged particles Why was the cold ethanol added to the soap and salt
mixture Describe the appearance of your final product Draw a diagram of DNA containing 5 sets of nucleotide
bases labelling the hydrogen bonds between the bases References and Resources Adapted from Berry Full
of DNA by Diane Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
53
HTTPWWWGENOMEBCCAEDUCATIONTEACHERSCLASSROOM-ACTIVITIESDNA-EXTRACTION
54
EXTRACHROMOSOMES EXIST IN PAIRS Because our chromosomes exist in pairs (and consequently we
have 2 alleles of each gene) we are a diploid species This is why our somatic cells are represented as 2n Our gametes (sperm and ova) on the other hand are haploid and are represented as n Other species may have different ploidy for example
triploid (3n) seedless watermelons tetaploid (4n) salmonidae fish pentaploid (5n) Kenai birch hexaploid (6n) some types of wheat kiwi fruit octaploid (8n) acipenser (a genus
of sturgeon fish strawberies) decaploid (10n) some strawberries dodecaploid (12n) some types of amphibians eg Xenopus
ruwenzoriensis
wwwcyberusca
55
YOUR EXTRACTED DNA UNDER THE MICROSCOPE LOOKED
LIKE
Collaboration-Brainstorming on what to look at under the
- DNA extraction- DNA is too small for even a microscope to see and after the extraction it just appears like a blob True or false
Could you any DNA strand
wwwemployeescsbsjuedu
56
HANDOUT ON DNA EXTRACTIONBACKGROUND EVERY DAY LIFE
EXTRA READING MATERIAL(EXTENSION )
DISCOVERING DNA DNA ON THE INSIDE USE OF DNA EXTRACTION INN EVERYDAY LIFE WHY IS DNA TESTING GOOD MOLECULAR BIOLOGY PCR-based diagnostics of genomic DNA
57
REFERENCES httpwwwsquidoocomhow-to-get-dna-from-a-kiwi-fruit Why Is DNA Testing Good | eHowcom
httpwwwehowcomabout_6684410_dna-testing-good_htmlixzz1MkbcIJs5 Why Is DNA Extraction Important | eHowcom
httpwwwehowcomlist_5839095_dna-extraction-important_htmlixzz1MkZDrqyY
Why Is Sodium Used in DNA Extraction | eHowcom httpwwwehowcomabout_6504902_sodium-used-dna-extraction_htmlixzz1MkaGWg57
Why Is Cold Alcohol Used in DNA Tests | eHowcom httpwwwehowcomabout_6399349_cold-alcohol-used-dna-tests_htmlixzz1MkbEmEdu
httplearngeneticsutaheducontentlabsextractionhowto www kamibudaksainsblogspotcom
httpwwwchemwatch goldcom References and Resources Adapted from Berry Full of DNA by Diane
Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
58
REFERENCES
Uses of DNA Extraction | eHowcom httpwwwehowcomabout_5344428_uses-dna-extractionhtmlixzz1MkWQB2TZ
Science 10 A Contextual Approach Heineman Queensland Science Projects Regan Spence Maggie Spenceley
httpenwikipediaorgwikiMolecular_biology httpwwwclinchemorgcgicontent
extract44102201 http
employeescsbsjueduhjakubowskiclassesch331dnaoldnastructurehtml
httpwwwgenomebccaeducationteachersclassroom-activitiesdna-extraction
12
ISOPROPANOL ndash MSDS HAND OUTS CHEMWATCH ISSUE DATE 15052010 ISOPROPANOL - FLAMMABLE Dangerous Good 3 SAFETY-DATA(tm) Ratings (Provided here for your convenience)
----------------------------------------------------------------------------------------------------------- Health Rating 2 - Moderate Flammability Rating 3 - Severe (Flammable) Reactivity Rating 2 - Moderate Contact Rating 3 - Severe Lab Protective Equip GOGGLES amp SHIELD LAB COAT amp APRON VENT HOOD PROPER GLOVES CLASS B EXTINGUISHER Storage Colour Code Red (Flammable)
Potential Health Effects ----------------------------------
Inhalation Inhalation of vapours irritates the respiratory tract Exposure to high concentrations has a narcotic effect producing symptoms of dizziness drowsiness headache staggering unconsciousness and possibly death Ingestion Can cause drowsiness unconsciousness and death Gastrointestinal pain cramps nausea vomiting and diarrhoea may also result The single lethal dose for a human adult = about 250 mls (8 ounces) Skin Contact May cause irritation with redness and pain May be absorbed through the skin with possible systemic effects Eye Contact Vapours cause eye irritation Splashes cause severe irritation possible corneal burns and eye damage Chronic Exposure Chronic exposure may cause skin effects Aggravation of Pre-existing Conditions Persons with pre-existing skin disorders or impaired liver kidney or pulmonary function may be more susceptible to the effects of this agent
13
METHYLENE BLUE ndash MSDS HAND OUTS CHEMWATCH ISSUE DATE 15052010 METHYLENE BLUE SAFETY-DATA(tm) Ratings (Provided here for your convenience)
----------------------------------------------------------------------------------------------------------- Health Rating 2 - Moderate Flammability Rating 1 - Slight Reactivity Rating 1 - Slight Contact Rating 1 - Slight Lab Protective Equip GOGGLES LAB COAT VENT HOOD PROPER GLOVES Storage Colour Code Green (General Storage) -----------------------------------------------------------------------------------------------------------
Potential Health Effects ----------------------------------
This material is relatively nonhazardous in routine industrial situations
Inhalation No adverse health effects expected from inhalation May cause a short period of rapid or difficult breathing Ingestion A burning sensation of the mouth may be noted following ingestion of methylene blue May cause nausea vomiting diarrhea and gastritis Large doses may cause abdominal and chest pain headache profuse sweating mental confusion painful maturation and methemoglobinemia Skin Contact Not expected to be a health hazard from skin exposure Methylene blue may colour the skin a bluish colour May cause photosensitization Eye Contact No adverse effects expected May cause mechanical irritation Chronic Exposure No information found Aggravation of Pre-existing Conditions No information found
14
NEW WORDS TO WORD BANKADD THESE TO YOUR EXCEL ALPHA TABLE
Precipitate Spooling Denaturising Lysing Homogenized Aggregate Lipid Centrifuging PCR
WORDS MEANING
15
bullDO YOU THINK YOU HAVE VERY MUCH IN COMMON WITH A KIWI FRUIT
BELIEVE IT OR NOT A KIWIS GENETIC MATERIAL IS VERY SIMILAR TO YOUR OWN
SEE AND TOUCH THE GENETIC MATERIAL THAT YOULL EXTRACT FROM THE CELLS OF A KIWI FRUIT
Activity Overview
Extract DNA from kiwi fruit using simple household chemicals
16
HT
TP
LEA
RN
GEN
ETIC
SU
TA
HE
DU
CO
NTE
NTL
AB
SE
XTR
AC
TIO
NH
OW
TO
DNA EXTRACTION USING KIWI FRUIT
17
AIM
TO
EX
TR
AC
T D
NA
FR
OM
TH
E C
ELLS
OF K
IWI F
RU
IT
wwwexploratoriumedu
MATERIALS EQUIPMENT (per student group takes 30 minutes)
frac12 cup Kiwi fruit 200 mL water dishwashing detergent dropping pipette fine mesh kitchen strainer amp cheese cloth glass rod large beaker large test tube light microscope meat tenderiser methylene blue microscope lamp inoculation needle microscope slide and cover slip paper towelling small beaker of alcohol (Isopropanol) spatula test-tube rack mortar and pestle
18
PART A EXTRACTING THE DNA
METHOD
1 Peal and Place the kiwi and water in the beaker and mush it with a mortar
and pestle DNA SOURCE About 125 ml 12 cup Twice as much water as DNA source (250
ml 1 cup) Table salt large pinch 1g 14 teaspoon Stir until the mixture is of a thin soupy
consistency
TAKE NOTES ndash NO TEXTBOOKS ALLOWED WHEN PERFORMING EXPERIMENT ndashMUST RECALL PROCEDURES METHOD
19
WHAT MUST I DO NOW
STRAIN THE DNA MIXTURE
DISHWASHING DETERGENT
20
2 Pour the thin cell mixture through the kitchen strainer (cheesecloth) into another large beaker
3 Measure 12 ml of the soup into
a small beakerAdd 2 ml of liquid detergentSwirl to mix stir thoroughly using a
glass rodLet mixture sit in container with
hot tap water (60 - 65degC) for 5 - 10 minutes
21
WHAT MUST I DO NOWMEAT TENDERIZER
ENZYME POWDERALCOHOL SEPARATION
wwwforumsoverclockerscomau
22
4 Add a pinch of enzyme (meat tenderizer) to mixture
(Using pineapple juice or contact lens cleaning solution will do the same as tenderizer)
Gently stir with toothpick skewer for 5 minutes continue stirring not too vigorously Be careful If you stir too hard youll break up the DNA making it harder to see
5 Quarter-fill a large test tube with the mixture
23
SLOWLY pour the same amount ice - cold (70 - 95 isopropyl or ethyl alcohol) into test tube pour alcohol down the side of the test tube
Tilting the test tube will make this easier to do
It forms a layer on top of the cell mixture
DO NOT MIX THE TWO LAYERS TOGETHER
Amount of alcohol and mixture should be the same
24
SEPARATION USING ALCOHOL
25
7 Observe the mixture for a few minutes You will see a white threadlike substance rise from the mixture to rest above in the alcohol layer
This is the DNA that you have extracted from the cells of the kiwi fruit
8 You can get more DNA to precipitate from the solution using a DNA collecting tool (glass or paper clip hook or cut inoculation needle)
Gently lift the water solution up into the alcohol layer (this allows more DNA to get in contact with the alcohol and precipitate)
26
INFORMATION DNA precipitates as a white stringy
ldquosnottyrdquo film at the water - alcohol interface and eventually will rise into the alcohol layer from the mixture layer
Allow test tube to sit for several minutes The clearer the DNA is the fewer
impurities you have
If you have an acceptable amount of DNA it can be spooled by rotating your collecting tool and then transferred into a clean tube container
27
WOW ndash SPOOLING THE DNA If you are careful you may be able wind up the DNA
around a glass rod or a skewer Position the tip of the glass rod or skewer where you can see the threads of DNA Steadily twist the rod or skewer as if you were making candy floss Alternatively use a straw to pull it out by suction ndash be careful not to get in you mouth
Donrsquot go too quickly
You should be able to pull the strands of DNA out of the mixture
ASK STUDENTS TO TAKE MOBILE PHONE PICTURES OF THEIR OWN EXTRACTED DNA AND COMPARE APPEARANCE WITH OTHERS
Photos of steps can be inserted in flow diagram(ICT)
28
KIWIDNA
29
If you want to save your DNA you can transfer it to a small container filled with alcohol
Leave tube container uncapped until the ethanol has evaporated
DNA can be stored in the fridge (dry or water buffer can be added)
You can now investigate the property
30
WHAT IS THAT STRINGY STUFFDNA IS A LONG STRINGY MOLECULE THE SALT THAT YOU ADDED IN STEP ONE HELPS IT STICK TOGETHER SO WHAT YOU SEE ARE CLUMPS OF TANGLED DNA MOLECULES
DNA NORMALLY STAYS DISSOLVED IN WATER BUT WHEN SALTY DNA COMES IN CONTACT WITH ALCOHOL IT BECOMES UNDISSOLVED THIS IS CALLED PRECIPITATION THE PHYSICAL FORCE OF THE DNA CLUMPING TOGETHER AS IT PRECIPITATES PULLS MORE STRANDS ALONG WITH IT AS IT RISES INTO THE ALCOHOL
31
THE WHYrsquoS Blending separated the pea cells In order to extract DNA from a cell the associated
membranes and proteins must first be removed (break apart the cells) and then physically separated (loosen the tough cell wall) from the DNA
To see the DNA we have to break open these two sacks We do this with detergent and salt( Sodium can be
involved in several of the steps ) Sodium is an element Its chemical symbol is Na for
Natrium the Latin word for sodium It is a positive ion and often associates with negative ions as part of useful compounds
Salt help precipitate protein and carbohydrates away from the DNA
Salt helps strip away the proteins associated with DNA + and ndash charge
32
THE WHYrsquoS Salt and Detergents are used to break down
cell walls and nuclear membranes to release the DNA
They work by chemically poking holes in the cell membranes or walls
Once holes are poked in the membranes the membranes can be further disrupted mechanically as with a blender
After that it is easier to get the contents of the cell out including the DNA
LETS HAVE A LOOK AT THE CELL
33
CELL TO DNA
httplearngeneticsutaheducontentlabsextractionhowtoHOW TO EXTRACT DNA FROM ANYTHING LIVING
34
THE WHYrsquoS WHY DETERGENT
Each cell is surrounded by a sack (the cell membrane)
DNA is found inside the second sack (nucleus) within each cell
To see the DNA we have to break it open A cell membrane has 2 layers of lipid(fat)
molecules with protein going through them When the lysis buffer (detergent) comes
close to the cell it captures the lipids and the proteins - breaks open the cell destroying the fatty membrane - DNA now released into the solution
35
A CELLS MEMBRANES HAVE TWO LAYERS OF LIPID (FAT) MOLECULES WITH PROTEINS GOING THROUGH THEM
36
Why detergent How does detergent work
Think about why you use soap to wash dishes or your hands To remove grease and dirt right
Soap molecules and grease molecules are made of two parts
1(Blue) Heads which like water 2(Green) Tails which hate water
37
AFTER ADDING THE DETERGENT WHAT DO YOU HAVE IN YOUR SOUP
38
THE WHYrsquoS ENZYME (MEAT TENDERIZER)
The DNA in the nucleus of the cell is molded folded and protected by proteins The tenderizer cut the proteins away from the DNA
In this experiment meat tenderizer acts as an enzyme to cut proteins just like a pair of scissors
The meat tenderizer cuts the proteins away from the DNA
39
THE WHYrsquoS CUTTING PROTEIN AND DNA
The DNA in the nucleus of the cell is moulded folded and protected by proteins
40
THE WHYrsquoSALCOHOL Alcohol is less dense than water so it floats
on top Look for clumps of white stringy stuff where the water and alcohol layers meet
DNA is not soluble in alcohol - other cell parts are
By adding alcohol DNA precipitates out of the solution and collect at the interface of the alcohol and soap layer
The colder the alcohol the less soluble the DNA will be in it
41
COLD ALCOHOL
DNA dissolves in water but precipitates in alcohol
Cold alcohol is used to separate DNA out of water-based solutions
This allows the DNA to be purified for subsequent genetic testing
Adding alcohol to a solution containing DNA is a simple way to obtain the pure DNA required and colder temperatures slow down enzymes that can break down DNA giving better extraction results
42
WHY IS DNA EXTRACTION IMPORTANT DNA extraction is an important molecular
biology procedure
By definition extraction is taking DNA out of any type of cell for the purpose of analysis
See Handouts for more information on the use of DNA extracted (extension only)
43
SUMMARIZEDNA EXTRACTING FROM KIWI FRUIT
44
BREAK DOWN CELL WALLS
Cells walls have to be destroyed to reach the DNA within cells
Extraction procedures to obtain pure DNA have to get rid of all the molecular and chemical components of the tissue from which the DNA is being extracted
First steps involve lysing or destroying the cell walls This can be done with a variety of chemical agents that are caustic to cell membranes but do not harm the DNA Cells can also be sonicated homogenized or ground up to destroy membranes
45
REMOVING Removing Lipids
Once the cell walls have been destroyed a detergent is added to get rid of the fats and oils that make up the cell membranes Detergents cause the fats and oils to dissolve into the solution
Remove ProteinsProteins and enzymes can be digested by
adding a protease to the solution Proteases break down proteins into small peptides and amino acids
46
PRECIPITATING AND PURIFYING Precipitate DNA
Adding cold alcohol to a solution will cause DNA to precipitate and aggregate The DNA can be collected by centrifuging the sample and pouring off the liquid layer The DNA should exist as a small pellet in the bottom of the centrifuge tube
Purify DNAThe DNA can be washed by re-suspending it
in a cold alcohol solution and re-centrifuging it several times to obtain a very pure DNA sample Typical alcohols used to precipitate DNA include ethanol and Isopropanol This process leaves a very pure sample for stringent DNA testing
47
PART B OBSERVING EXTRACTED DNA UNDER THE MICROSCOPE
48
PART B A CLOSER LOOK
8 Use a dropping pipette to carefully remove some of the threadlike substance from the top of your preparation
9 Place one or two drops onto the middle of a
microscope slide
10 Add two drops of methylene blue Wait 3 or 4 minutes to allow the methylene blue to be absorbed by the DNA
11 Carefully place a cover slip on the slide Gently press
a folded piece of paper towelling over the top of the prepared slide to soak up any excess liquid
12 Observe the DNA under low power then high power
49
DISCUSSIONANSWER IN GROUP RELATED WORKED 1 Write a detailed description of the material
floating at the top of the test tube after the alcohol was added
2 Describe the DNA as it appears under the
microscope under high power 3 Prepare a diagram of your DNA specimen 4 What was the reason for using methylene
blue
50
HOMEWORKCONSTRUCT A FLOW CHART DRAWING MIND MAP OF THE PROCESS YOU USED EXTRACTING THE DNA FROM KIWIrsquoS
Next to each step explain how the method you used was important (Hint What substances make up the membranes of cells and cell organelles
How do detergents work
What is the active ingredient in meat tenderiser
51
TIPS STEPS FOR FLOW CHART In order to release the DNA from the
nuclei of the pea cells you first separated the cells from one another
Then the cell and nuclear membranes needed to be ruptured to release the cell contents and the contents of the nucleus
Once removed from the nuclear membrane the DNA had to be untangled into the visible threadlike structures you ended up with
52
HOMEWORK EXTENSION GROUP QUESTIONS Where can DNA be found in the cell Discuss the action of the soap (detergent) on the cell
What is the purpose of the soap in this activity What was the purpose of the Sodium Chloride Include a
discussion of polarity and charged particles Why was the cold ethanol added to the soap and salt
mixture Describe the appearance of your final product Draw a diagram of DNA containing 5 sets of nucleotide
bases labelling the hydrogen bonds between the bases References and Resources Adapted from Berry Full
of DNA by Diane Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
53
HTTPWWWGENOMEBCCAEDUCATIONTEACHERSCLASSROOM-ACTIVITIESDNA-EXTRACTION
54
EXTRACHROMOSOMES EXIST IN PAIRS Because our chromosomes exist in pairs (and consequently we
have 2 alleles of each gene) we are a diploid species This is why our somatic cells are represented as 2n Our gametes (sperm and ova) on the other hand are haploid and are represented as n Other species may have different ploidy for example
triploid (3n) seedless watermelons tetaploid (4n) salmonidae fish pentaploid (5n) Kenai birch hexaploid (6n) some types of wheat kiwi fruit octaploid (8n) acipenser (a genus
of sturgeon fish strawberies) decaploid (10n) some strawberries dodecaploid (12n) some types of amphibians eg Xenopus
ruwenzoriensis
wwwcyberusca
55
YOUR EXTRACTED DNA UNDER THE MICROSCOPE LOOKED
LIKE
Collaboration-Brainstorming on what to look at under the
- DNA extraction- DNA is too small for even a microscope to see and after the extraction it just appears like a blob True or false
Could you any DNA strand
wwwemployeescsbsjuedu
56
HANDOUT ON DNA EXTRACTIONBACKGROUND EVERY DAY LIFE
EXTRA READING MATERIAL(EXTENSION )
DISCOVERING DNA DNA ON THE INSIDE USE OF DNA EXTRACTION INN EVERYDAY LIFE WHY IS DNA TESTING GOOD MOLECULAR BIOLOGY PCR-based diagnostics of genomic DNA
57
REFERENCES httpwwwsquidoocomhow-to-get-dna-from-a-kiwi-fruit Why Is DNA Testing Good | eHowcom
httpwwwehowcomabout_6684410_dna-testing-good_htmlixzz1MkbcIJs5 Why Is DNA Extraction Important | eHowcom
httpwwwehowcomlist_5839095_dna-extraction-important_htmlixzz1MkZDrqyY
Why Is Sodium Used in DNA Extraction | eHowcom httpwwwehowcomabout_6504902_sodium-used-dna-extraction_htmlixzz1MkaGWg57
Why Is Cold Alcohol Used in DNA Tests | eHowcom httpwwwehowcomabout_6399349_cold-alcohol-used-dna-tests_htmlixzz1MkbEmEdu
httplearngeneticsutaheducontentlabsextractionhowto www kamibudaksainsblogspotcom
httpwwwchemwatch goldcom References and Resources Adapted from Berry Full of DNA by Diane
Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
58
REFERENCES
Uses of DNA Extraction | eHowcom httpwwwehowcomabout_5344428_uses-dna-extractionhtmlixzz1MkWQB2TZ
Science 10 A Contextual Approach Heineman Queensland Science Projects Regan Spence Maggie Spenceley
httpenwikipediaorgwikiMolecular_biology httpwwwclinchemorgcgicontent
extract44102201 http
employeescsbsjueduhjakubowskiclassesch331dnaoldnastructurehtml
httpwwwgenomebccaeducationteachersclassroom-activitiesdna-extraction
13
METHYLENE BLUE ndash MSDS HAND OUTS CHEMWATCH ISSUE DATE 15052010 METHYLENE BLUE SAFETY-DATA(tm) Ratings (Provided here for your convenience)
----------------------------------------------------------------------------------------------------------- Health Rating 2 - Moderate Flammability Rating 1 - Slight Reactivity Rating 1 - Slight Contact Rating 1 - Slight Lab Protective Equip GOGGLES LAB COAT VENT HOOD PROPER GLOVES Storage Colour Code Green (General Storage) -----------------------------------------------------------------------------------------------------------
Potential Health Effects ----------------------------------
This material is relatively nonhazardous in routine industrial situations
Inhalation No adverse health effects expected from inhalation May cause a short period of rapid or difficult breathing Ingestion A burning sensation of the mouth may be noted following ingestion of methylene blue May cause nausea vomiting diarrhea and gastritis Large doses may cause abdominal and chest pain headache profuse sweating mental confusion painful maturation and methemoglobinemia Skin Contact Not expected to be a health hazard from skin exposure Methylene blue may colour the skin a bluish colour May cause photosensitization Eye Contact No adverse effects expected May cause mechanical irritation Chronic Exposure No information found Aggravation of Pre-existing Conditions No information found
14
NEW WORDS TO WORD BANKADD THESE TO YOUR EXCEL ALPHA TABLE
Precipitate Spooling Denaturising Lysing Homogenized Aggregate Lipid Centrifuging PCR
WORDS MEANING
15
bullDO YOU THINK YOU HAVE VERY MUCH IN COMMON WITH A KIWI FRUIT
BELIEVE IT OR NOT A KIWIS GENETIC MATERIAL IS VERY SIMILAR TO YOUR OWN
SEE AND TOUCH THE GENETIC MATERIAL THAT YOULL EXTRACT FROM THE CELLS OF A KIWI FRUIT
Activity Overview
Extract DNA from kiwi fruit using simple household chemicals
16
HT
TP
LEA
RN
GEN
ETIC
SU
TA
HE
DU
CO
NTE
NTL
AB
SE
XTR
AC
TIO
NH
OW
TO
DNA EXTRACTION USING KIWI FRUIT
17
AIM
TO
EX
TR
AC
T D
NA
FR
OM
TH
E C
ELLS
OF K
IWI F
RU
IT
wwwexploratoriumedu
MATERIALS EQUIPMENT (per student group takes 30 minutes)
frac12 cup Kiwi fruit 200 mL water dishwashing detergent dropping pipette fine mesh kitchen strainer amp cheese cloth glass rod large beaker large test tube light microscope meat tenderiser methylene blue microscope lamp inoculation needle microscope slide and cover slip paper towelling small beaker of alcohol (Isopropanol) spatula test-tube rack mortar and pestle
18
PART A EXTRACTING THE DNA
METHOD
1 Peal and Place the kiwi and water in the beaker and mush it with a mortar
and pestle DNA SOURCE About 125 ml 12 cup Twice as much water as DNA source (250
ml 1 cup) Table salt large pinch 1g 14 teaspoon Stir until the mixture is of a thin soupy
consistency
TAKE NOTES ndash NO TEXTBOOKS ALLOWED WHEN PERFORMING EXPERIMENT ndashMUST RECALL PROCEDURES METHOD
19
WHAT MUST I DO NOW
STRAIN THE DNA MIXTURE
DISHWASHING DETERGENT
20
2 Pour the thin cell mixture through the kitchen strainer (cheesecloth) into another large beaker
3 Measure 12 ml of the soup into
a small beakerAdd 2 ml of liquid detergentSwirl to mix stir thoroughly using a
glass rodLet mixture sit in container with
hot tap water (60 - 65degC) for 5 - 10 minutes
21
WHAT MUST I DO NOWMEAT TENDERIZER
ENZYME POWDERALCOHOL SEPARATION
wwwforumsoverclockerscomau
22
4 Add a pinch of enzyme (meat tenderizer) to mixture
(Using pineapple juice or contact lens cleaning solution will do the same as tenderizer)
Gently stir with toothpick skewer for 5 minutes continue stirring not too vigorously Be careful If you stir too hard youll break up the DNA making it harder to see
5 Quarter-fill a large test tube with the mixture
23
SLOWLY pour the same amount ice - cold (70 - 95 isopropyl or ethyl alcohol) into test tube pour alcohol down the side of the test tube
Tilting the test tube will make this easier to do
It forms a layer on top of the cell mixture
DO NOT MIX THE TWO LAYERS TOGETHER
Amount of alcohol and mixture should be the same
24
SEPARATION USING ALCOHOL
25
7 Observe the mixture for a few minutes You will see a white threadlike substance rise from the mixture to rest above in the alcohol layer
This is the DNA that you have extracted from the cells of the kiwi fruit
8 You can get more DNA to precipitate from the solution using a DNA collecting tool (glass or paper clip hook or cut inoculation needle)
Gently lift the water solution up into the alcohol layer (this allows more DNA to get in contact with the alcohol and precipitate)
26
INFORMATION DNA precipitates as a white stringy
ldquosnottyrdquo film at the water - alcohol interface and eventually will rise into the alcohol layer from the mixture layer
Allow test tube to sit for several minutes The clearer the DNA is the fewer
impurities you have
If you have an acceptable amount of DNA it can be spooled by rotating your collecting tool and then transferred into a clean tube container
27
WOW ndash SPOOLING THE DNA If you are careful you may be able wind up the DNA
around a glass rod or a skewer Position the tip of the glass rod or skewer where you can see the threads of DNA Steadily twist the rod or skewer as if you were making candy floss Alternatively use a straw to pull it out by suction ndash be careful not to get in you mouth
Donrsquot go too quickly
You should be able to pull the strands of DNA out of the mixture
ASK STUDENTS TO TAKE MOBILE PHONE PICTURES OF THEIR OWN EXTRACTED DNA AND COMPARE APPEARANCE WITH OTHERS
Photos of steps can be inserted in flow diagram(ICT)
28
KIWIDNA
29
If you want to save your DNA you can transfer it to a small container filled with alcohol
Leave tube container uncapped until the ethanol has evaporated
DNA can be stored in the fridge (dry or water buffer can be added)
You can now investigate the property
30
WHAT IS THAT STRINGY STUFFDNA IS A LONG STRINGY MOLECULE THE SALT THAT YOU ADDED IN STEP ONE HELPS IT STICK TOGETHER SO WHAT YOU SEE ARE CLUMPS OF TANGLED DNA MOLECULES
DNA NORMALLY STAYS DISSOLVED IN WATER BUT WHEN SALTY DNA COMES IN CONTACT WITH ALCOHOL IT BECOMES UNDISSOLVED THIS IS CALLED PRECIPITATION THE PHYSICAL FORCE OF THE DNA CLUMPING TOGETHER AS IT PRECIPITATES PULLS MORE STRANDS ALONG WITH IT AS IT RISES INTO THE ALCOHOL
31
THE WHYrsquoS Blending separated the pea cells In order to extract DNA from a cell the associated
membranes and proteins must first be removed (break apart the cells) and then physically separated (loosen the tough cell wall) from the DNA
To see the DNA we have to break open these two sacks We do this with detergent and salt( Sodium can be
involved in several of the steps ) Sodium is an element Its chemical symbol is Na for
Natrium the Latin word for sodium It is a positive ion and often associates with negative ions as part of useful compounds
Salt help precipitate protein and carbohydrates away from the DNA
Salt helps strip away the proteins associated with DNA + and ndash charge
32
THE WHYrsquoS Salt and Detergents are used to break down
cell walls and nuclear membranes to release the DNA
They work by chemically poking holes in the cell membranes or walls
Once holes are poked in the membranes the membranes can be further disrupted mechanically as with a blender
After that it is easier to get the contents of the cell out including the DNA
LETS HAVE A LOOK AT THE CELL
33
CELL TO DNA
httplearngeneticsutaheducontentlabsextractionhowtoHOW TO EXTRACT DNA FROM ANYTHING LIVING
34
THE WHYrsquoS WHY DETERGENT
Each cell is surrounded by a sack (the cell membrane)
DNA is found inside the second sack (nucleus) within each cell
To see the DNA we have to break it open A cell membrane has 2 layers of lipid(fat)
molecules with protein going through them When the lysis buffer (detergent) comes
close to the cell it captures the lipids and the proteins - breaks open the cell destroying the fatty membrane - DNA now released into the solution
35
A CELLS MEMBRANES HAVE TWO LAYERS OF LIPID (FAT) MOLECULES WITH PROTEINS GOING THROUGH THEM
36
Why detergent How does detergent work
Think about why you use soap to wash dishes or your hands To remove grease and dirt right
Soap molecules and grease molecules are made of two parts
1(Blue) Heads which like water 2(Green) Tails which hate water
37
AFTER ADDING THE DETERGENT WHAT DO YOU HAVE IN YOUR SOUP
38
THE WHYrsquoS ENZYME (MEAT TENDERIZER)
The DNA in the nucleus of the cell is molded folded and protected by proteins The tenderizer cut the proteins away from the DNA
In this experiment meat tenderizer acts as an enzyme to cut proteins just like a pair of scissors
The meat tenderizer cuts the proteins away from the DNA
39
THE WHYrsquoS CUTTING PROTEIN AND DNA
The DNA in the nucleus of the cell is moulded folded and protected by proteins
40
THE WHYrsquoSALCOHOL Alcohol is less dense than water so it floats
on top Look for clumps of white stringy stuff where the water and alcohol layers meet
DNA is not soluble in alcohol - other cell parts are
By adding alcohol DNA precipitates out of the solution and collect at the interface of the alcohol and soap layer
The colder the alcohol the less soluble the DNA will be in it
41
COLD ALCOHOL
DNA dissolves in water but precipitates in alcohol
Cold alcohol is used to separate DNA out of water-based solutions
This allows the DNA to be purified for subsequent genetic testing
Adding alcohol to a solution containing DNA is a simple way to obtain the pure DNA required and colder temperatures slow down enzymes that can break down DNA giving better extraction results
42
WHY IS DNA EXTRACTION IMPORTANT DNA extraction is an important molecular
biology procedure
By definition extraction is taking DNA out of any type of cell for the purpose of analysis
See Handouts for more information on the use of DNA extracted (extension only)
43
SUMMARIZEDNA EXTRACTING FROM KIWI FRUIT
44
BREAK DOWN CELL WALLS
Cells walls have to be destroyed to reach the DNA within cells
Extraction procedures to obtain pure DNA have to get rid of all the molecular and chemical components of the tissue from which the DNA is being extracted
First steps involve lysing or destroying the cell walls This can be done with a variety of chemical agents that are caustic to cell membranes but do not harm the DNA Cells can also be sonicated homogenized or ground up to destroy membranes
45
REMOVING Removing Lipids
Once the cell walls have been destroyed a detergent is added to get rid of the fats and oils that make up the cell membranes Detergents cause the fats and oils to dissolve into the solution
Remove ProteinsProteins and enzymes can be digested by
adding a protease to the solution Proteases break down proteins into small peptides and amino acids
46
PRECIPITATING AND PURIFYING Precipitate DNA
Adding cold alcohol to a solution will cause DNA to precipitate and aggregate The DNA can be collected by centrifuging the sample and pouring off the liquid layer The DNA should exist as a small pellet in the bottom of the centrifuge tube
Purify DNAThe DNA can be washed by re-suspending it
in a cold alcohol solution and re-centrifuging it several times to obtain a very pure DNA sample Typical alcohols used to precipitate DNA include ethanol and Isopropanol This process leaves a very pure sample for stringent DNA testing
47
PART B OBSERVING EXTRACTED DNA UNDER THE MICROSCOPE
48
PART B A CLOSER LOOK
8 Use a dropping pipette to carefully remove some of the threadlike substance from the top of your preparation
9 Place one or two drops onto the middle of a
microscope slide
10 Add two drops of methylene blue Wait 3 or 4 minutes to allow the methylene blue to be absorbed by the DNA
11 Carefully place a cover slip on the slide Gently press
a folded piece of paper towelling over the top of the prepared slide to soak up any excess liquid
12 Observe the DNA under low power then high power
49
DISCUSSIONANSWER IN GROUP RELATED WORKED 1 Write a detailed description of the material
floating at the top of the test tube after the alcohol was added
2 Describe the DNA as it appears under the
microscope under high power 3 Prepare a diagram of your DNA specimen 4 What was the reason for using methylene
blue
50
HOMEWORKCONSTRUCT A FLOW CHART DRAWING MIND MAP OF THE PROCESS YOU USED EXTRACTING THE DNA FROM KIWIrsquoS
Next to each step explain how the method you used was important (Hint What substances make up the membranes of cells and cell organelles
How do detergents work
What is the active ingredient in meat tenderiser
51
TIPS STEPS FOR FLOW CHART In order to release the DNA from the
nuclei of the pea cells you first separated the cells from one another
Then the cell and nuclear membranes needed to be ruptured to release the cell contents and the contents of the nucleus
Once removed from the nuclear membrane the DNA had to be untangled into the visible threadlike structures you ended up with
52
HOMEWORK EXTENSION GROUP QUESTIONS Where can DNA be found in the cell Discuss the action of the soap (detergent) on the cell
What is the purpose of the soap in this activity What was the purpose of the Sodium Chloride Include a
discussion of polarity and charged particles Why was the cold ethanol added to the soap and salt
mixture Describe the appearance of your final product Draw a diagram of DNA containing 5 sets of nucleotide
bases labelling the hydrogen bonds between the bases References and Resources Adapted from Berry Full
of DNA by Diane Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
53
HTTPWWWGENOMEBCCAEDUCATIONTEACHERSCLASSROOM-ACTIVITIESDNA-EXTRACTION
54
EXTRACHROMOSOMES EXIST IN PAIRS Because our chromosomes exist in pairs (and consequently we
have 2 alleles of each gene) we are a diploid species This is why our somatic cells are represented as 2n Our gametes (sperm and ova) on the other hand are haploid and are represented as n Other species may have different ploidy for example
triploid (3n) seedless watermelons tetaploid (4n) salmonidae fish pentaploid (5n) Kenai birch hexaploid (6n) some types of wheat kiwi fruit octaploid (8n) acipenser (a genus
of sturgeon fish strawberies) decaploid (10n) some strawberries dodecaploid (12n) some types of amphibians eg Xenopus
ruwenzoriensis
wwwcyberusca
55
YOUR EXTRACTED DNA UNDER THE MICROSCOPE LOOKED
LIKE
Collaboration-Brainstorming on what to look at under the
- DNA extraction- DNA is too small for even a microscope to see and after the extraction it just appears like a blob True or false
Could you any DNA strand
wwwemployeescsbsjuedu
56
HANDOUT ON DNA EXTRACTIONBACKGROUND EVERY DAY LIFE
EXTRA READING MATERIAL(EXTENSION )
DISCOVERING DNA DNA ON THE INSIDE USE OF DNA EXTRACTION INN EVERYDAY LIFE WHY IS DNA TESTING GOOD MOLECULAR BIOLOGY PCR-based diagnostics of genomic DNA
57
REFERENCES httpwwwsquidoocomhow-to-get-dna-from-a-kiwi-fruit Why Is DNA Testing Good | eHowcom
httpwwwehowcomabout_6684410_dna-testing-good_htmlixzz1MkbcIJs5 Why Is DNA Extraction Important | eHowcom
httpwwwehowcomlist_5839095_dna-extraction-important_htmlixzz1MkZDrqyY
Why Is Sodium Used in DNA Extraction | eHowcom httpwwwehowcomabout_6504902_sodium-used-dna-extraction_htmlixzz1MkaGWg57
Why Is Cold Alcohol Used in DNA Tests | eHowcom httpwwwehowcomabout_6399349_cold-alcohol-used-dna-tests_htmlixzz1MkbEmEdu
httplearngeneticsutaheducontentlabsextractionhowto www kamibudaksainsblogspotcom
httpwwwchemwatch goldcom References and Resources Adapted from Berry Full of DNA by Diane
Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
58
REFERENCES
Uses of DNA Extraction | eHowcom httpwwwehowcomabout_5344428_uses-dna-extractionhtmlixzz1MkWQB2TZ
Science 10 A Contextual Approach Heineman Queensland Science Projects Regan Spence Maggie Spenceley
httpenwikipediaorgwikiMolecular_biology httpwwwclinchemorgcgicontent
extract44102201 http
employeescsbsjueduhjakubowskiclassesch331dnaoldnastructurehtml
httpwwwgenomebccaeducationteachersclassroom-activitiesdna-extraction
14
NEW WORDS TO WORD BANKADD THESE TO YOUR EXCEL ALPHA TABLE
Precipitate Spooling Denaturising Lysing Homogenized Aggregate Lipid Centrifuging PCR
WORDS MEANING
15
bullDO YOU THINK YOU HAVE VERY MUCH IN COMMON WITH A KIWI FRUIT
BELIEVE IT OR NOT A KIWIS GENETIC MATERIAL IS VERY SIMILAR TO YOUR OWN
SEE AND TOUCH THE GENETIC MATERIAL THAT YOULL EXTRACT FROM THE CELLS OF A KIWI FRUIT
Activity Overview
Extract DNA from kiwi fruit using simple household chemicals
16
HT
TP
LEA
RN
GEN
ETIC
SU
TA
HE
DU
CO
NTE
NTL
AB
SE
XTR
AC
TIO
NH
OW
TO
DNA EXTRACTION USING KIWI FRUIT
17
AIM
TO
EX
TR
AC
T D
NA
FR
OM
TH
E C
ELLS
OF K
IWI F
RU
IT
wwwexploratoriumedu
MATERIALS EQUIPMENT (per student group takes 30 minutes)
frac12 cup Kiwi fruit 200 mL water dishwashing detergent dropping pipette fine mesh kitchen strainer amp cheese cloth glass rod large beaker large test tube light microscope meat tenderiser methylene blue microscope lamp inoculation needle microscope slide and cover slip paper towelling small beaker of alcohol (Isopropanol) spatula test-tube rack mortar and pestle
18
PART A EXTRACTING THE DNA
METHOD
1 Peal and Place the kiwi and water in the beaker and mush it with a mortar
and pestle DNA SOURCE About 125 ml 12 cup Twice as much water as DNA source (250
ml 1 cup) Table salt large pinch 1g 14 teaspoon Stir until the mixture is of a thin soupy
consistency
TAKE NOTES ndash NO TEXTBOOKS ALLOWED WHEN PERFORMING EXPERIMENT ndashMUST RECALL PROCEDURES METHOD
19
WHAT MUST I DO NOW
STRAIN THE DNA MIXTURE
DISHWASHING DETERGENT
20
2 Pour the thin cell mixture through the kitchen strainer (cheesecloth) into another large beaker
3 Measure 12 ml of the soup into
a small beakerAdd 2 ml of liquid detergentSwirl to mix stir thoroughly using a
glass rodLet mixture sit in container with
hot tap water (60 - 65degC) for 5 - 10 minutes
21
WHAT MUST I DO NOWMEAT TENDERIZER
ENZYME POWDERALCOHOL SEPARATION
wwwforumsoverclockerscomau
22
4 Add a pinch of enzyme (meat tenderizer) to mixture
(Using pineapple juice or contact lens cleaning solution will do the same as tenderizer)
Gently stir with toothpick skewer for 5 minutes continue stirring not too vigorously Be careful If you stir too hard youll break up the DNA making it harder to see
5 Quarter-fill a large test tube with the mixture
23
SLOWLY pour the same amount ice - cold (70 - 95 isopropyl or ethyl alcohol) into test tube pour alcohol down the side of the test tube
Tilting the test tube will make this easier to do
It forms a layer on top of the cell mixture
DO NOT MIX THE TWO LAYERS TOGETHER
Amount of alcohol and mixture should be the same
24
SEPARATION USING ALCOHOL
25
7 Observe the mixture for a few minutes You will see a white threadlike substance rise from the mixture to rest above in the alcohol layer
This is the DNA that you have extracted from the cells of the kiwi fruit
8 You can get more DNA to precipitate from the solution using a DNA collecting tool (glass or paper clip hook or cut inoculation needle)
Gently lift the water solution up into the alcohol layer (this allows more DNA to get in contact with the alcohol and precipitate)
26
INFORMATION DNA precipitates as a white stringy
ldquosnottyrdquo film at the water - alcohol interface and eventually will rise into the alcohol layer from the mixture layer
Allow test tube to sit for several minutes The clearer the DNA is the fewer
impurities you have
If you have an acceptable amount of DNA it can be spooled by rotating your collecting tool and then transferred into a clean tube container
27
WOW ndash SPOOLING THE DNA If you are careful you may be able wind up the DNA
around a glass rod or a skewer Position the tip of the glass rod or skewer where you can see the threads of DNA Steadily twist the rod or skewer as if you were making candy floss Alternatively use a straw to pull it out by suction ndash be careful not to get in you mouth
Donrsquot go too quickly
You should be able to pull the strands of DNA out of the mixture
ASK STUDENTS TO TAKE MOBILE PHONE PICTURES OF THEIR OWN EXTRACTED DNA AND COMPARE APPEARANCE WITH OTHERS
Photos of steps can be inserted in flow diagram(ICT)
28
KIWIDNA
29
If you want to save your DNA you can transfer it to a small container filled with alcohol
Leave tube container uncapped until the ethanol has evaporated
DNA can be stored in the fridge (dry or water buffer can be added)
You can now investigate the property
30
WHAT IS THAT STRINGY STUFFDNA IS A LONG STRINGY MOLECULE THE SALT THAT YOU ADDED IN STEP ONE HELPS IT STICK TOGETHER SO WHAT YOU SEE ARE CLUMPS OF TANGLED DNA MOLECULES
DNA NORMALLY STAYS DISSOLVED IN WATER BUT WHEN SALTY DNA COMES IN CONTACT WITH ALCOHOL IT BECOMES UNDISSOLVED THIS IS CALLED PRECIPITATION THE PHYSICAL FORCE OF THE DNA CLUMPING TOGETHER AS IT PRECIPITATES PULLS MORE STRANDS ALONG WITH IT AS IT RISES INTO THE ALCOHOL
31
THE WHYrsquoS Blending separated the pea cells In order to extract DNA from a cell the associated
membranes and proteins must first be removed (break apart the cells) and then physically separated (loosen the tough cell wall) from the DNA
To see the DNA we have to break open these two sacks We do this with detergent and salt( Sodium can be
involved in several of the steps ) Sodium is an element Its chemical symbol is Na for
Natrium the Latin word for sodium It is a positive ion and often associates with negative ions as part of useful compounds
Salt help precipitate protein and carbohydrates away from the DNA
Salt helps strip away the proteins associated with DNA + and ndash charge
32
THE WHYrsquoS Salt and Detergents are used to break down
cell walls and nuclear membranes to release the DNA
They work by chemically poking holes in the cell membranes or walls
Once holes are poked in the membranes the membranes can be further disrupted mechanically as with a blender
After that it is easier to get the contents of the cell out including the DNA
LETS HAVE A LOOK AT THE CELL
33
CELL TO DNA
httplearngeneticsutaheducontentlabsextractionhowtoHOW TO EXTRACT DNA FROM ANYTHING LIVING
34
THE WHYrsquoS WHY DETERGENT
Each cell is surrounded by a sack (the cell membrane)
DNA is found inside the second sack (nucleus) within each cell
To see the DNA we have to break it open A cell membrane has 2 layers of lipid(fat)
molecules with protein going through them When the lysis buffer (detergent) comes
close to the cell it captures the lipids and the proteins - breaks open the cell destroying the fatty membrane - DNA now released into the solution
35
A CELLS MEMBRANES HAVE TWO LAYERS OF LIPID (FAT) MOLECULES WITH PROTEINS GOING THROUGH THEM
36
Why detergent How does detergent work
Think about why you use soap to wash dishes or your hands To remove grease and dirt right
Soap molecules and grease molecules are made of two parts
1(Blue) Heads which like water 2(Green) Tails which hate water
37
AFTER ADDING THE DETERGENT WHAT DO YOU HAVE IN YOUR SOUP
38
THE WHYrsquoS ENZYME (MEAT TENDERIZER)
The DNA in the nucleus of the cell is molded folded and protected by proteins The tenderizer cut the proteins away from the DNA
In this experiment meat tenderizer acts as an enzyme to cut proteins just like a pair of scissors
The meat tenderizer cuts the proteins away from the DNA
39
THE WHYrsquoS CUTTING PROTEIN AND DNA
The DNA in the nucleus of the cell is moulded folded and protected by proteins
40
THE WHYrsquoSALCOHOL Alcohol is less dense than water so it floats
on top Look for clumps of white stringy stuff where the water and alcohol layers meet
DNA is not soluble in alcohol - other cell parts are
By adding alcohol DNA precipitates out of the solution and collect at the interface of the alcohol and soap layer
The colder the alcohol the less soluble the DNA will be in it
41
COLD ALCOHOL
DNA dissolves in water but precipitates in alcohol
Cold alcohol is used to separate DNA out of water-based solutions
This allows the DNA to be purified for subsequent genetic testing
Adding alcohol to a solution containing DNA is a simple way to obtain the pure DNA required and colder temperatures slow down enzymes that can break down DNA giving better extraction results
42
WHY IS DNA EXTRACTION IMPORTANT DNA extraction is an important molecular
biology procedure
By definition extraction is taking DNA out of any type of cell for the purpose of analysis
See Handouts for more information on the use of DNA extracted (extension only)
43
SUMMARIZEDNA EXTRACTING FROM KIWI FRUIT
44
BREAK DOWN CELL WALLS
Cells walls have to be destroyed to reach the DNA within cells
Extraction procedures to obtain pure DNA have to get rid of all the molecular and chemical components of the tissue from which the DNA is being extracted
First steps involve lysing or destroying the cell walls This can be done with a variety of chemical agents that are caustic to cell membranes but do not harm the DNA Cells can also be sonicated homogenized or ground up to destroy membranes
45
REMOVING Removing Lipids
Once the cell walls have been destroyed a detergent is added to get rid of the fats and oils that make up the cell membranes Detergents cause the fats and oils to dissolve into the solution
Remove ProteinsProteins and enzymes can be digested by
adding a protease to the solution Proteases break down proteins into small peptides and amino acids
46
PRECIPITATING AND PURIFYING Precipitate DNA
Adding cold alcohol to a solution will cause DNA to precipitate and aggregate The DNA can be collected by centrifuging the sample and pouring off the liquid layer The DNA should exist as a small pellet in the bottom of the centrifuge tube
Purify DNAThe DNA can be washed by re-suspending it
in a cold alcohol solution and re-centrifuging it several times to obtain a very pure DNA sample Typical alcohols used to precipitate DNA include ethanol and Isopropanol This process leaves a very pure sample for stringent DNA testing
47
PART B OBSERVING EXTRACTED DNA UNDER THE MICROSCOPE
48
PART B A CLOSER LOOK
8 Use a dropping pipette to carefully remove some of the threadlike substance from the top of your preparation
9 Place one or two drops onto the middle of a
microscope slide
10 Add two drops of methylene blue Wait 3 or 4 minutes to allow the methylene blue to be absorbed by the DNA
11 Carefully place a cover slip on the slide Gently press
a folded piece of paper towelling over the top of the prepared slide to soak up any excess liquid
12 Observe the DNA under low power then high power
49
DISCUSSIONANSWER IN GROUP RELATED WORKED 1 Write a detailed description of the material
floating at the top of the test tube after the alcohol was added
2 Describe the DNA as it appears under the
microscope under high power 3 Prepare a diagram of your DNA specimen 4 What was the reason for using methylene
blue
50
HOMEWORKCONSTRUCT A FLOW CHART DRAWING MIND MAP OF THE PROCESS YOU USED EXTRACTING THE DNA FROM KIWIrsquoS
Next to each step explain how the method you used was important (Hint What substances make up the membranes of cells and cell organelles
How do detergents work
What is the active ingredient in meat tenderiser
51
TIPS STEPS FOR FLOW CHART In order to release the DNA from the
nuclei of the pea cells you first separated the cells from one another
Then the cell and nuclear membranes needed to be ruptured to release the cell contents and the contents of the nucleus
Once removed from the nuclear membrane the DNA had to be untangled into the visible threadlike structures you ended up with
52
HOMEWORK EXTENSION GROUP QUESTIONS Where can DNA be found in the cell Discuss the action of the soap (detergent) on the cell
What is the purpose of the soap in this activity What was the purpose of the Sodium Chloride Include a
discussion of polarity and charged particles Why was the cold ethanol added to the soap and salt
mixture Describe the appearance of your final product Draw a diagram of DNA containing 5 sets of nucleotide
bases labelling the hydrogen bonds between the bases References and Resources Adapted from Berry Full
of DNA by Diane Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
53
HTTPWWWGENOMEBCCAEDUCATIONTEACHERSCLASSROOM-ACTIVITIESDNA-EXTRACTION
54
EXTRACHROMOSOMES EXIST IN PAIRS Because our chromosomes exist in pairs (and consequently we
have 2 alleles of each gene) we are a diploid species This is why our somatic cells are represented as 2n Our gametes (sperm and ova) on the other hand are haploid and are represented as n Other species may have different ploidy for example
triploid (3n) seedless watermelons tetaploid (4n) salmonidae fish pentaploid (5n) Kenai birch hexaploid (6n) some types of wheat kiwi fruit octaploid (8n) acipenser (a genus
of sturgeon fish strawberies) decaploid (10n) some strawberries dodecaploid (12n) some types of amphibians eg Xenopus
ruwenzoriensis
wwwcyberusca
55
YOUR EXTRACTED DNA UNDER THE MICROSCOPE LOOKED
LIKE
Collaboration-Brainstorming on what to look at under the
- DNA extraction- DNA is too small for even a microscope to see and after the extraction it just appears like a blob True or false
Could you any DNA strand
wwwemployeescsbsjuedu
56
HANDOUT ON DNA EXTRACTIONBACKGROUND EVERY DAY LIFE
EXTRA READING MATERIAL(EXTENSION )
DISCOVERING DNA DNA ON THE INSIDE USE OF DNA EXTRACTION INN EVERYDAY LIFE WHY IS DNA TESTING GOOD MOLECULAR BIOLOGY PCR-based diagnostics of genomic DNA
57
REFERENCES httpwwwsquidoocomhow-to-get-dna-from-a-kiwi-fruit Why Is DNA Testing Good | eHowcom
httpwwwehowcomabout_6684410_dna-testing-good_htmlixzz1MkbcIJs5 Why Is DNA Extraction Important | eHowcom
httpwwwehowcomlist_5839095_dna-extraction-important_htmlixzz1MkZDrqyY
Why Is Sodium Used in DNA Extraction | eHowcom httpwwwehowcomabout_6504902_sodium-used-dna-extraction_htmlixzz1MkaGWg57
Why Is Cold Alcohol Used in DNA Tests | eHowcom httpwwwehowcomabout_6399349_cold-alcohol-used-dna-tests_htmlixzz1MkbEmEdu
httplearngeneticsutaheducontentlabsextractionhowto www kamibudaksainsblogspotcom
httpwwwchemwatch goldcom References and Resources Adapted from Berry Full of DNA by Diane
Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
58
REFERENCES
Uses of DNA Extraction | eHowcom httpwwwehowcomabout_5344428_uses-dna-extractionhtmlixzz1MkWQB2TZ
Science 10 A Contextual Approach Heineman Queensland Science Projects Regan Spence Maggie Spenceley
httpenwikipediaorgwikiMolecular_biology httpwwwclinchemorgcgicontent
extract44102201 http
employeescsbsjueduhjakubowskiclassesch331dnaoldnastructurehtml
httpwwwgenomebccaeducationteachersclassroom-activitiesdna-extraction
15
bullDO YOU THINK YOU HAVE VERY MUCH IN COMMON WITH A KIWI FRUIT
BELIEVE IT OR NOT A KIWIS GENETIC MATERIAL IS VERY SIMILAR TO YOUR OWN
SEE AND TOUCH THE GENETIC MATERIAL THAT YOULL EXTRACT FROM THE CELLS OF A KIWI FRUIT
Activity Overview
Extract DNA from kiwi fruit using simple household chemicals
16
HT
TP
LEA
RN
GEN
ETIC
SU
TA
HE
DU
CO
NTE
NTL
AB
SE
XTR
AC
TIO
NH
OW
TO
DNA EXTRACTION USING KIWI FRUIT
17
AIM
TO
EX
TR
AC
T D
NA
FR
OM
TH
E C
ELLS
OF K
IWI F
RU
IT
wwwexploratoriumedu
MATERIALS EQUIPMENT (per student group takes 30 minutes)
frac12 cup Kiwi fruit 200 mL water dishwashing detergent dropping pipette fine mesh kitchen strainer amp cheese cloth glass rod large beaker large test tube light microscope meat tenderiser methylene blue microscope lamp inoculation needle microscope slide and cover slip paper towelling small beaker of alcohol (Isopropanol) spatula test-tube rack mortar and pestle
18
PART A EXTRACTING THE DNA
METHOD
1 Peal and Place the kiwi and water in the beaker and mush it with a mortar
and pestle DNA SOURCE About 125 ml 12 cup Twice as much water as DNA source (250
ml 1 cup) Table salt large pinch 1g 14 teaspoon Stir until the mixture is of a thin soupy
consistency
TAKE NOTES ndash NO TEXTBOOKS ALLOWED WHEN PERFORMING EXPERIMENT ndashMUST RECALL PROCEDURES METHOD
19
WHAT MUST I DO NOW
STRAIN THE DNA MIXTURE
DISHWASHING DETERGENT
20
2 Pour the thin cell mixture through the kitchen strainer (cheesecloth) into another large beaker
3 Measure 12 ml of the soup into
a small beakerAdd 2 ml of liquid detergentSwirl to mix stir thoroughly using a
glass rodLet mixture sit in container with
hot tap water (60 - 65degC) for 5 - 10 minutes
21
WHAT MUST I DO NOWMEAT TENDERIZER
ENZYME POWDERALCOHOL SEPARATION
wwwforumsoverclockerscomau
22
4 Add a pinch of enzyme (meat tenderizer) to mixture
(Using pineapple juice or contact lens cleaning solution will do the same as tenderizer)
Gently stir with toothpick skewer for 5 minutes continue stirring not too vigorously Be careful If you stir too hard youll break up the DNA making it harder to see
5 Quarter-fill a large test tube with the mixture
23
SLOWLY pour the same amount ice - cold (70 - 95 isopropyl or ethyl alcohol) into test tube pour alcohol down the side of the test tube
Tilting the test tube will make this easier to do
It forms a layer on top of the cell mixture
DO NOT MIX THE TWO LAYERS TOGETHER
Amount of alcohol and mixture should be the same
24
SEPARATION USING ALCOHOL
25
7 Observe the mixture for a few minutes You will see a white threadlike substance rise from the mixture to rest above in the alcohol layer
This is the DNA that you have extracted from the cells of the kiwi fruit
8 You can get more DNA to precipitate from the solution using a DNA collecting tool (glass or paper clip hook or cut inoculation needle)
Gently lift the water solution up into the alcohol layer (this allows more DNA to get in contact with the alcohol and precipitate)
26
INFORMATION DNA precipitates as a white stringy
ldquosnottyrdquo film at the water - alcohol interface and eventually will rise into the alcohol layer from the mixture layer
Allow test tube to sit for several minutes The clearer the DNA is the fewer
impurities you have
If you have an acceptable amount of DNA it can be spooled by rotating your collecting tool and then transferred into a clean tube container
27
WOW ndash SPOOLING THE DNA If you are careful you may be able wind up the DNA
around a glass rod or a skewer Position the tip of the glass rod or skewer where you can see the threads of DNA Steadily twist the rod or skewer as if you were making candy floss Alternatively use a straw to pull it out by suction ndash be careful not to get in you mouth
Donrsquot go too quickly
You should be able to pull the strands of DNA out of the mixture
ASK STUDENTS TO TAKE MOBILE PHONE PICTURES OF THEIR OWN EXTRACTED DNA AND COMPARE APPEARANCE WITH OTHERS
Photos of steps can be inserted in flow diagram(ICT)
28
KIWIDNA
29
If you want to save your DNA you can transfer it to a small container filled with alcohol
Leave tube container uncapped until the ethanol has evaporated
DNA can be stored in the fridge (dry or water buffer can be added)
You can now investigate the property
30
WHAT IS THAT STRINGY STUFFDNA IS A LONG STRINGY MOLECULE THE SALT THAT YOU ADDED IN STEP ONE HELPS IT STICK TOGETHER SO WHAT YOU SEE ARE CLUMPS OF TANGLED DNA MOLECULES
DNA NORMALLY STAYS DISSOLVED IN WATER BUT WHEN SALTY DNA COMES IN CONTACT WITH ALCOHOL IT BECOMES UNDISSOLVED THIS IS CALLED PRECIPITATION THE PHYSICAL FORCE OF THE DNA CLUMPING TOGETHER AS IT PRECIPITATES PULLS MORE STRANDS ALONG WITH IT AS IT RISES INTO THE ALCOHOL
31
THE WHYrsquoS Blending separated the pea cells In order to extract DNA from a cell the associated
membranes and proteins must first be removed (break apart the cells) and then physically separated (loosen the tough cell wall) from the DNA
To see the DNA we have to break open these two sacks We do this with detergent and salt( Sodium can be
involved in several of the steps ) Sodium is an element Its chemical symbol is Na for
Natrium the Latin word for sodium It is a positive ion and often associates with negative ions as part of useful compounds
Salt help precipitate protein and carbohydrates away from the DNA
Salt helps strip away the proteins associated with DNA + and ndash charge
32
THE WHYrsquoS Salt and Detergents are used to break down
cell walls and nuclear membranes to release the DNA
They work by chemically poking holes in the cell membranes or walls
Once holes are poked in the membranes the membranes can be further disrupted mechanically as with a blender
After that it is easier to get the contents of the cell out including the DNA
LETS HAVE A LOOK AT THE CELL
33
CELL TO DNA
httplearngeneticsutaheducontentlabsextractionhowtoHOW TO EXTRACT DNA FROM ANYTHING LIVING
34
THE WHYrsquoS WHY DETERGENT
Each cell is surrounded by a sack (the cell membrane)
DNA is found inside the second sack (nucleus) within each cell
To see the DNA we have to break it open A cell membrane has 2 layers of lipid(fat)
molecules with protein going through them When the lysis buffer (detergent) comes
close to the cell it captures the lipids and the proteins - breaks open the cell destroying the fatty membrane - DNA now released into the solution
35
A CELLS MEMBRANES HAVE TWO LAYERS OF LIPID (FAT) MOLECULES WITH PROTEINS GOING THROUGH THEM
36
Why detergent How does detergent work
Think about why you use soap to wash dishes or your hands To remove grease and dirt right
Soap molecules and grease molecules are made of two parts
1(Blue) Heads which like water 2(Green) Tails which hate water
37
AFTER ADDING THE DETERGENT WHAT DO YOU HAVE IN YOUR SOUP
38
THE WHYrsquoS ENZYME (MEAT TENDERIZER)
The DNA in the nucleus of the cell is molded folded and protected by proteins The tenderizer cut the proteins away from the DNA
In this experiment meat tenderizer acts as an enzyme to cut proteins just like a pair of scissors
The meat tenderizer cuts the proteins away from the DNA
39
THE WHYrsquoS CUTTING PROTEIN AND DNA
The DNA in the nucleus of the cell is moulded folded and protected by proteins
40
THE WHYrsquoSALCOHOL Alcohol is less dense than water so it floats
on top Look for clumps of white stringy stuff where the water and alcohol layers meet
DNA is not soluble in alcohol - other cell parts are
By adding alcohol DNA precipitates out of the solution and collect at the interface of the alcohol and soap layer
The colder the alcohol the less soluble the DNA will be in it
41
COLD ALCOHOL
DNA dissolves in water but precipitates in alcohol
Cold alcohol is used to separate DNA out of water-based solutions
This allows the DNA to be purified for subsequent genetic testing
Adding alcohol to a solution containing DNA is a simple way to obtain the pure DNA required and colder temperatures slow down enzymes that can break down DNA giving better extraction results
42
WHY IS DNA EXTRACTION IMPORTANT DNA extraction is an important molecular
biology procedure
By definition extraction is taking DNA out of any type of cell for the purpose of analysis
See Handouts for more information on the use of DNA extracted (extension only)
43
SUMMARIZEDNA EXTRACTING FROM KIWI FRUIT
44
BREAK DOWN CELL WALLS
Cells walls have to be destroyed to reach the DNA within cells
Extraction procedures to obtain pure DNA have to get rid of all the molecular and chemical components of the tissue from which the DNA is being extracted
First steps involve lysing or destroying the cell walls This can be done with a variety of chemical agents that are caustic to cell membranes but do not harm the DNA Cells can also be sonicated homogenized or ground up to destroy membranes
45
REMOVING Removing Lipids
Once the cell walls have been destroyed a detergent is added to get rid of the fats and oils that make up the cell membranes Detergents cause the fats and oils to dissolve into the solution
Remove ProteinsProteins and enzymes can be digested by
adding a protease to the solution Proteases break down proteins into small peptides and amino acids
46
PRECIPITATING AND PURIFYING Precipitate DNA
Adding cold alcohol to a solution will cause DNA to precipitate and aggregate The DNA can be collected by centrifuging the sample and pouring off the liquid layer The DNA should exist as a small pellet in the bottom of the centrifuge tube
Purify DNAThe DNA can be washed by re-suspending it
in a cold alcohol solution and re-centrifuging it several times to obtain a very pure DNA sample Typical alcohols used to precipitate DNA include ethanol and Isopropanol This process leaves a very pure sample for stringent DNA testing
47
PART B OBSERVING EXTRACTED DNA UNDER THE MICROSCOPE
48
PART B A CLOSER LOOK
8 Use a dropping pipette to carefully remove some of the threadlike substance from the top of your preparation
9 Place one or two drops onto the middle of a
microscope slide
10 Add two drops of methylene blue Wait 3 or 4 minutes to allow the methylene blue to be absorbed by the DNA
11 Carefully place a cover slip on the slide Gently press
a folded piece of paper towelling over the top of the prepared slide to soak up any excess liquid
12 Observe the DNA under low power then high power
49
DISCUSSIONANSWER IN GROUP RELATED WORKED 1 Write a detailed description of the material
floating at the top of the test tube after the alcohol was added
2 Describe the DNA as it appears under the
microscope under high power 3 Prepare a diagram of your DNA specimen 4 What was the reason for using methylene
blue
50
HOMEWORKCONSTRUCT A FLOW CHART DRAWING MIND MAP OF THE PROCESS YOU USED EXTRACTING THE DNA FROM KIWIrsquoS
Next to each step explain how the method you used was important (Hint What substances make up the membranes of cells and cell organelles
How do detergents work
What is the active ingredient in meat tenderiser
51
TIPS STEPS FOR FLOW CHART In order to release the DNA from the
nuclei of the pea cells you first separated the cells from one another
Then the cell and nuclear membranes needed to be ruptured to release the cell contents and the contents of the nucleus
Once removed from the nuclear membrane the DNA had to be untangled into the visible threadlike structures you ended up with
52
HOMEWORK EXTENSION GROUP QUESTIONS Where can DNA be found in the cell Discuss the action of the soap (detergent) on the cell
What is the purpose of the soap in this activity What was the purpose of the Sodium Chloride Include a
discussion of polarity and charged particles Why was the cold ethanol added to the soap and salt
mixture Describe the appearance of your final product Draw a diagram of DNA containing 5 sets of nucleotide
bases labelling the hydrogen bonds between the bases References and Resources Adapted from Berry Full
of DNA by Diane Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
53
HTTPWWWGENOMEBCCAEDUCATIONTEACHERSCLASSROOM-ACTIVITIESDNA-EXTRACTION
54
EXTRACHROMOSOMES EXIST IN PAIRS Because our chromosomes exist in pairs (and consequently we
have 2 alleles of each gene) we are a diploid species This is why our somatic cells are represented as 2n Our gametes (sperm and ova) on the other hand are haploid and are represented as n Other species may have different ploidy for example
triploid (3n) seedless watermelons tetaploid (4n) salmonidae fish pentaploid (5n) Kenai birch hexaploid (6n) some types of wheat kiwi fruit octaploid (8n) acipenser (a genus
of sturgeon fish strawberies) decaploid (10n) some strawberries dodecaploid (12n) some types of amphibians eg Xenopus
ruwenzoriensis
wwwcyberusca
55
YOUR EXTRACTED DNA UNDER THE MICROSCOPE LOOKED
LIKE
Collaboration-Brainstorming on what to look at under the
- DNA extraction- DNA is too small for even a microscope to see and after the extraction it just appears like a blob True or false
Could you any DNA strand
wwwemployeescsbsjuedu
56
HANDOUT ON DNA EXTRACTIONBACKGROUND EVERY DAY LIFE
EXTRA READING MATERIAL(EXTENSION )
DISCOVERING DNA DNA ON THE INSIDE USE OF DNA EXTRACTION INN EVERYDAY LIFE WHY IS DNA TESTING GOOD MOLECULAR BIOLOGY PCR-based diagnostics of genomic DNA
57
REFERENCES httpwwwsquidoocomhow-to-get-dna-from-a-kiwi-fruit Why Is DNA Testing Good | eHowcom
httpwwwehowcomabout_6684410_dna-testing-good_htmlixzz1MkbcIJs5 Why Is DNA Extraction Important | eHowcom
httpwwwehowcomlist_5839095_dna-extraction-important_htmlixzz1MkZDrqyY
Why Is Sodium Used in DNA Extraction | eHowcom httpwwwehowcomabout_6504902_sodium-used-dna-extraction_htmlixzz1MkaGWg57
Why Is Cold Alcohol Used in DNA Tests | eHowcom httpwwwehowcomabout_6399349_cold-alcohol-used-dna-tests_htmlixzz1MkbEmEdu
httplearngeneticsutaheducontentlabsextractionhowto www kamibudaksainsblogspotcom
httpwwwchemwatch goldcom References and Resources Adapted from Berry Full of DNA by Diane
Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
58
REFERENCES
Uses of DNA Extraction | eHowcom httpwwwehowcomabout_5344428_uses-dna-extractionhtmlixzz1MkWQB2TZ
Science 10 A Contextual Approach Heineman Queensland Science Projects Regan Spence Maggie Spenceley
httpenwikipediaorgwikiMolecular_biology httpwwwclinchemorgcgicontent
extract44102201 http
employeescsbsjueduhjakubowskiclassesch331dnaoldnastructurehtml
httpwwwgenomebccaeducationteachersclassroom-activitiesdna-extraction
16
HT
TP
LEA
RN
GEN
ETIC
SU
TA
HE
DU
CO
NTE
NTL
AB
SE
XTR
AC
TIO
NH
OW
TO
DNA EXTRACTION USING KIWI FRUIT
17
AIM
TO
EX
TR
AC
T D
NA
FR
OM
TH
E C
ELLS
OF K
IWI F
RU
IT
wwwexploratoriumedu
MATERIALS EQUIPMENT (per student group takes 30 minutes)
frac12 cup Kiwi fruit 200 mL water dishwashing detergent dropping pipette fine mesh kitchen strainer amp cheese cloth glass rod large beaker large test tube light microscope meat tenderiser methylene blue microscope lamp inoculation needle microscope slide and cover slip paper towelling small beaker of alcohol (Isopropanol) spatula test-tube rack mortar and pestle
18
PART A EXTRACTING THE DNA
METHOD
1 Peal and Place the kiwi and water in the beaker and mush it with a mortar
and pestle DNA SOURCE About 125 ml 12 cup Twice as much water as DNA source (250
ml 1 cup) Table salt large pinch 1g 14 teaspoon Stir until the mixture is of a thin soupy
consistency
TAKE NOTES ndash NO TEXTBOOKS ALLOWED WHEN PERFORMING EXPERIMENT ndashMUST RECALL PROCEDURES METHOD
19
WHAT MUST I DO NOW
STRAIN THE DNA MIXTURE
DISHWASHING DETERGENT
20
2 Pour the thin cell mixture through the kitchen strainer (cheesecloth) into another large beaker
3 Measure 12 ml of the soup into
a small beakerAdd 2 ml of liquid detergentSwirl to mix stir thoroughly using a
glass rodLet mixture sit in container with
hot tap water (60 - 65degC) for 5 - 10 minutes
21
WHAT MUST I DO NOWMEAT TENDERIZER
ENZYME POWDERALCOHOL SEPARATION
wwwforumsoverclockerscomau
22
4 Add a pinch of enzyme (meat tenderizer) to mixture
(Using pineapple juice or contact lens cleaning solution will do the same as tenderizer)
Gently stir with toothpick skewer for 5 minutes continue stirring not too vigorously Be careful If you stir too hard youll break up the DNA making it harder to see
5 Quarter-fill a large test tube with the mixture
23
SLOWLY pour the same amount ice - cold (70 - 95 isopropyl or ethyl alcohol) into test tube pour alcohol down the side of the test tube
Tilting the test tube will make this easier to do
It forms a layer on top of the cell mixture
DO NOT MIX THE TWO LAYERS TOGETHER
Amount of alcohol and mixture should be the same
24
SEPARATION USING ALCOHOL
25
7 Observe the mixture for a few minutes You will see a white threadlike substance rise from the mixture to rest above in the alcohol layer
This is the DNA that you have extracted from the cells of the kiwi fruit
8 You can get more DNA to precipitate from the solution using a DNA collecting tool (glass or paper clip hook or cut inoculation needle)
Gently lift the water solution up into the alcohol layer (this allows more DNA to get in contact with the alcohol and precipitate)
26
INFORMATION DNA precipitates as a white stringy
ldquosnottyrdquo film at the water - alcohol interface and eventually will rise into the alcohol layer from the mixture layer
Allow test tube to sit for several minutes The clearer the DNA is the fewer
impurities you have
If you have an acceptable amount of DNA it can be spooled by rotating your collecting tool and then transferred into a clean tube container
27
WOW ndash SPOOLING THE DNA If you are careful you may be able wind up the DNA
around a glass rod or a skewer Position the tip of the glass rod or skewer where you can see the threads of DNA Steadily twist the rod or skewer as if you were making candy floss Alternatively use a straw to pull it out by suction ndash be careful not to get in you mouth
Donrsquot go too quickly
You should be able to pull the strands of DNA out of the mixture
ASK STUDENTS TO TAKE MOBILE PHONE PICTURES OF THEIR OWN EXTRACTED DNA AND COMPARE APPEARANCE WITH OTHERS
Photos of steps can be inserted in flow diagram(ICT)
28
KIWIDNA
29
If you want to save your DNA you can transfer it to a small container filled with alcohol
Leave tube container uncapped until the ethanol has evaporated
DNA can be stored in the fridge (dry or water buffer can be added)
You can now investigate the property
30
WHAT IS THAT STRINGY STUFFDNA IS A LONG STRINGY MOLECULE THE SALT THAT YOU ADDED IN STEP ONE HELPS IT STICK TOGETHER SO WHAT YOU SEE ARE CLUMPS OF TANGLED DNA MOLECULES
DNA NORMALLY STAYS DISSOLVED IN WATER BUT WHEN SALTY DNA COMES IN CONTACT WITH ALCOHOL IT BECOMES UNDISSOLVED THIS IS CALLED PRECIPITATION THE PHYSICAL FORCE OF THE DNA CLUMPING TOGETHER AS IT PRECIPITATES PULLS MORE STRANDS ALONG WITH IT AS IT RISES INTO THE ALCOHOL
31
THE WHYrsquoS Blending separated the pea cells In order to extract DNA from a cell the associated
membranes and proteins must first be removed (break apart the cells) and then physically separated (loosen the tough cell wall) from the DNA
To see the DNA we have to break open these two sacks We do this with detergent and salt( Sodium can be
involved in several of the steps ) Sodium is an element Its chemical symbol is Na for
Natrium the Latin word for sodium It is a positive ion and often associates with negative ions as part of useful compounds
Salt help precipitate protein and carbohydrates away from the DNA
Salt helps strip away the proteins associated with DNA + and ndash charge
32
THE WHYrsquoS Salt and Detergents are used to break down
cell walls and nuclear membranes to release the DNA
They work by chemically poking holes in the cell membranes or walls
Once holes are poked in the membranes the membranes can be further disrupted mechanically as with a blender
After that it is easier to get the contents of the cell out including the DNA
LETS HAVE A LOOK AT THE CELL
33
CELL TO DNA
httplearngeneticsutaheducontentlabsextractionhowtoHOW TO EXTRACT DNA FROM ANYTHING LIVING
34
THE WHYrsquoS WHY DETERGENT
Each cell is surrounded by a sack (the cell membrane)
DNA is found inside the second sack (nucleus) within each cell
To see the DNA we have to break it open A cell membrane has 2 layers of lipid(fat)
molecules with protein going through them When the lysis buffer (detergent) comes
close to the cell it captures the lipids and the proteins - breaks open the cell destroying the fatty membrane - DNA now released into the solution
35
A CELLS MEMBRANES HAVE TWO LAYERS OF LIPID (FAT) MOLECULES WITH PROTEINS GOING THROUGH THEM
36
Why detergent How does detergent work
Think about why you use soap to wash dishes or your hands To remove grease and dirt right
Soap molecules and grease molecules are made of two parts
1(Blue) Heads which like water 2(Green) Tails which hate water
37
AFTER ADDING THE DETERGENT WHAT DO YOU HAVE IN YOUR SOUP
38
THE WHYrsquoS ENZYME (MEAT TENDERIZER)
The DNA in the nucleus of the cell is molded folded and protected by proteins The tenderizer cut the proteins away from the DNA
In this experiment meat tenderizer acts as an enzyme to cut proteins just like a pair of scissors
The meat tenderizer cuts the proteins away from the DNA
39
THE WHYrsquoS CUTTING PROTEIN AND DNA
The DNA in the nucleus of the cell is moulded folded and protected by proteins
40
THE WHYrsquoSALCOHOL Alcohol is less dense than water so it floats
on top Look for clumps of white stringy stuff where the water and alcohol layers meet
DNA is not soluble in alcohol - other cell parts are
By adding alcohol DNA precipitates out of the solution and collect at the interface of the alcohol and soap layer
The colder the alcohol the less soluble the DNA will be in it
41
COLD ALCOHOL
DNA dissolves in water but precipitates in alcohol
Cold alcohol is used to separate DNA out of water-based solutions
This allows the DNA to be purified for subsequent genetic testing
Adding alcohol to a solution containing DNA is a simple way to obtain the pure DNA required and colder temperatures slow down enzymes that can break down DNA giving better extraction results
42
WHY IS DNA EXTRACTION IMPORTANT DNA extraction is an important molecular
biology procedure
By definition extraction is taking DNA out of any type of cell for the purpose of analysis
See Handouts for more information on the use of DNA extracted (extension only)
43
SUMMARIZEDNA EXTRACTING FROM KIWI FRUIT
44
BREAK DOWN CELL WALLS
Cells walls have to be destroyed to reach the DNA within cells
Extraction procedures to obtain pure DNA have to get rid of all the molecular and chemical components of the tissue from which the DNA is being extracted
First steps involve lysing or destroying the cell walls This can be done with a variety of chemical agents that are caustic to cell membranes but do not harm the DNA Cells can also be sonicated homogenized or ground up to destroy membranes
45
REMOVING Removing Lipids
Once the cell walls have been destroyed a detergent is added to get rid of the fats and oils that make up the cell membranes Detergents cause the fats and oils to dissolve into the solution
Remove ProteinsProteins and enzymes can be digested by
adding a protease to the solution Proteases break down proteins into small peptides and amino acids
46
PRECIPITATING AND PURIFYING Precipitate DNA
Adding cold alcohol to a solution will cause DNA to precipitate and aggregate The DNA can be collected by centrifuging the sample and pouring off the liquid layer The DNA should exist as a small pellet in the bottom of the centrifuge tube
Purify DNAThe DNA can be washed by re-suspending it
in a cold alcohol solution and re-centrifuging it several times to obtain a very pure DNA sample Typical alcohols used to precipitate DNA include ethanol and Isopropanol This process leaves a very pure sample for stringent DNA testing
47
PART B OBSERVING EXTRACTED DNA UNDER THE MICROSCOPE
48
PART B A CLOSER LOOK
8 Use a dropping pipette to carefully remove some of the threadlike substance from the top of your preparation
9 Place one or two drops onto the middle of a
microscope slide
10 Add two drops of methylene blue Wait 3 or 4 minutes to allow the methylene blue to be absorbed by the DNA
11 Carefully place a cover slip on the slide Gently press
a folded piece of paper towelling over the top of the prepared slide to soak up any excess liquid
12 Observe the DNA under low power then high power
49
DISCUSSIONANSWER IN GROUP RELATED WORKED 1 Write a detailed description of the material
floating at the top of the test tube after the alcohol was added
2 Describe the DNA as it appears under the
microscope under high power 3 Prepare a diagram of your DNA specimen 4 What was the reason for using methylene
blue
50
HOMEWORKCONSTRUCT A FLOW CHART DRAWING MIND MAP OF THE PROCESS YOU USED EXTRACTING THE DNA FROM KIWIrsquoS
Next to each step explain how the method you used was important (Hint What substances make up the membranes of cells and cell organelles
How do detergents work
What is the active ingredient in meat tenderiser
51
TIPS STEPS FOR FLOW CHART In order to release the DNA from the
nuclei of the pea cells you first separated the cells from one another
Then the cell and nuclear membranes needed to be ruptured to release the cell contents and the contents of the nucleus
Once removed from the nuclear membrane the DNA had to be untangled into the visible threadlike structures you ended up with
52
HOMEWORK EXTENSION GROUP QUESTIONS Where can DNA be found in the cell Discuss the action of the soap (detergent) on the cell
What is the purpose of the soap in this activity What was the purpose of the Sodium Chloride Include a
discussion of polarity and charged particles Why was the cold ethanol added to the soap and salt
mixture Describe the appearance of your final product Draw a diagram of DNA containing 5 sets of nucleotide
bases labelling the hydrogen bonds between the bases References and Resources Adapted from Berry Full
of DNA by Diane Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
53
HTTPWWWGENOMEBCCAEDUCATIONTEACHERSCLASSROOM-ACTIVITIESDNA-EXTRACTION
54
EXTRACHROMOSOMES EXIST IN PAIRS Because our chromosomes exist in pairs (and consequently we
have 2 alleles of each gene) we are a diploid species This is why our somatic cells are represented as 2n Our gametes (sperm and ova) on the other hand are haploid and are represented as n Other species may have different ploidy for example
triploid (3n) seedless watermelons tetaploid (4n) salmonidae fish pentaploid (5n) Kenai birch hexaploid (6n) some types of wheat kiwi fruit octaploid (8n) acipenser (a genus
of sturgeon fish strawberies) decaploid (10n) some strawberries dodecaploid (12n) some types of amphibians eg Xenopus
ruwenzoriensis
wwwcyberusca
55
YOUR EXTRACTED DNA UNDER THE MICROSCOPE LOOKED
LIKE
Collaboration-Brainstorming on what to look at under the
- DNA extraction- DNA is too small for even a microscope to see and after the extraction it just appears like a blob True or false
Could you any DNA strand
wwwemployeescsbsjuedu
56
HANDOUT ON DNA EXTRACTIONBACKGROUND EVERY DAY LIFE
EXTRA READING MATERIAL(EXTENSION )
DISCOVERING DNA DNA ON THE INSIDE USE OF DNA EXTRACTION INN EVERYDAY LIFE WHY IS DNA TESTING GOOD MOLECULAR BIOLOGY PCR-based diagnostics of genomic DNA
57
REFERENCES httpwwwsquidoocomhow-to-get-dna-from-a-kiwi-fruit Why Is DNA Testing Good | eHowcom
httpwwwehowcomabout_6684410_dna-testing-good_htmlixzz1MkbcIJs5 Why Is DNA Extraction Important | eHowcom
httpwwwehowcomlist_5839095_dna-extraction-important_htmlixzz1MkZDrqyY
Why Is Sodium Used in DNA Extraction | eHowcom httpwwwehowcomabout_6504902_sodium-used-dna-extraction_htmlixzz1MkaGWg57
Why Is Cold Alcohol Used in DNA Tests | eHowcom httpwwwehowcomabout_6399349_cold-alcohol-used-dna-tests_htmlixzz1MkbEmEdu
httplearngeneticsutaheducontentlabsextractionhowto www kamibudaksainsblogspotcom
httpwwwchemwatch goldcom References and Resources Adapted from Berry Full of DNA by Diane
Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
58
REFERENCES
Uses of DNA Extraction | eHowcom httpwwwehowcomabout_5344428_uses-dna-extractionhtmlixzz1MkWQB2TZ
Science 10 A Contextual Approach Heineman Queensland Science Projects Regan Spence Maggie Spenceley
httpenwikipediaorgwikiMolecular_biology httpwwwclinchemorgcgicontent
extract44102201 http
employeescsbsjueduhjakubowskiclassesch331dnaoldnastructurehtml
httpwwwgenomebccaeducationteachersclassroom-activitiesdna-extraction
17
AIM
TO
EX
TR
AC
T D
NA
FR
OM
TH
E C
ELLS
OF K
IWI F
RU
IT
wwwexploratoriumedu
MATERIALS EQUIPMENT (per student group takes 30 minutes)
frac12 cup Kiwi fruit 200 mL water dishwashing detergent dropping pipette fine mesh kitchen strainer amp cheese cloth glass rod large beaker large test tube light microscope meat tenderiser methylene blue microscope lamp inoculation needle microscope slide and cover slip paper towelling small beaker of alcohol (Isopropanol) spatula test-tube rack mortar and pestle
18
PART A EXTRACTING THE DNA
METHOD
1 Peal and Place the kiwi and water in the beaker and mush it with a mortar
and pestle DNA SOURCE About 125 ml 12 cup Twice as much water as DNA source (250
ml 1 cup) Table salt large pinch 1g 14 teaspoon Stir until the mixture is of a thin soupy
consistency
TAKE NOTES ndash NO TEXTBOOKS ALLOWED WHEN PERFORMING EXPERIMENT ndashMUST RECALL PROCEDURES METHOD
19
WHAT MUST I DO NOW
STRAIN THE DNA MIXTURE
DISHWASHING DETERGENT
20
2 Pour the thin cell mixture through the kitchen strainer (cheesecloth) into another large beaker
3 Measure 12 ml of the soup into
a small beakerAdd 2 ml of liquid detergentSwirl to mix stir thoroughly using a
glass rodLet mixture sit in container with
hot tap water (60 - 65degC) for 5 - 10 minutes
21
WHAT MUST I DO NOWMEAT TENDERIZER
ENZYME POWDERALCOHOL SEPARATION
wwwforumsoverclockerscomau
22
4 Add a pinch of enzyme (meat tenderizer) to mixture
(Using pineapple juice or contact lens cleaning solution will do the same as tenderizer)
Gently stir with toothpick skewer for 5 minutes continue stirring not too vigorously Be careful If you stir too hard youll break up the DNA making it harder to see
5 Quarter-fill a large test tube with the mixture
23
SLOWLY pour the same amount ice - cold (70 - 95 isopropyl or ethyl alcohol) into test tube pour alcohol down the side of the test tube
Tilting the test tube will make this easier to do
It forms a layer on top of the cell mixture
DO NOT MIX THE TWO LAYERS TOGETHER
Amount of alcohol and mixture should be the same
24
SEPARATION USING ALCOHOL
25
7 Observe the mixture for a few minutes You will see a white threadlike substance rise from the mixture to rest above in the alcohol layer
This is the DNA that you have extracted from the cells of the kiwi fruit
8 You can get more DNA to precipitate from the solution using a DNA collecting tool (glass or paper clip hook or cut inoculation needle)
Gently lift the water solution up into the alcohol layer (this allows more DNA to get in contact with the alcohol and precipitate)
26
INFORMATION DNA precipitates as a white stringy
ldquosnottyrdquo film at the water - alcohol interface and eventually will rise into the alcohol layer from the mixture layer
Allow test tube to sit for several minutes The clearer the DNA is the fewer
impurities you have
If you have an acceptable amount of DNA it can be spooled by rotating your collecting tool and then transferred into a clean tube container
27
WOW ndash SPOOLING THE DNA If you are careful you may be able wind up the DNA
around a glass rod or a skewer Position the tip of the glass rod or skewer where you can see the threads of DNA Steadily twist the rod or skewer as if you were making candy floss Alternatively use a straw to pull it out by suction ndash be careful not to get in you mouth
Donrsquot go too quickly
You should be able to pull the strands of DNA out of the mixture
ASK STUDENTS TO TAKE MOBILE PHONE PICTURES OF THEIR OWN EXTRACTED DNA AND COMPARE APPEARANCE WITH OTHERS
Photos of steps can be inserted in flow diagram(ICT)
28
KIWIDNA
29
If you want to save your DNA you can transfer it to a small container filled with alcohol
Leave tube container uncapped until the ethanol has evaporated
DNA can be stored in the fridge (dry or water buffer can be added)
You can now investigate the property
30
WHAT IS THAT STRINGY STUFFDNA IS A LONG STRINGY MOLECULE THE SALT THAT YOU ADDED IN STEP ONE HELPS IT STICK TOGETHER SO WHAT YOU SEE ARE CLUMPS OF TANGLED DNA MOLECULES
DNA NORMALLY STAYS DISSOLVED IN WATER BUT WHEN SALTY DNA COMES IN CONTACT WITH ALCOHOL IT BECOMES UNDISSOLVED THIS IS CALLED PRECIPITATION THE PHYSICAL FORCE OF THE DNA CLUMPING TOGETHER AS IT PRECIPITATES PULLS MORE STRANDS ALONG WITH IT AS IT RISES INTO THE ALCOHOL
31
THE WHYrsquoS Blending separated the pea cells In order to extract DNA from a cell the associated
membranes and proteins must first be removed (break apart the cells) and then physically separated (loosen the tough cell wall) from the DNA
To see the DNA we have to break open these two sacks We do this with detergent and salt( Sodium can be
involved in several of the steps ) Sodium is an element Its chemical symbol is Na for
Natrium the Latin word for sodium It is a positive ion and often associates with negative ions as part of useful compounds
Salt help precipitate protein and carbohydrates away from the DNA
Salt helps strip away the proteins associated with DNA + and ndash charge
32
THE WHYrsquoS Salt and Detergents are used to break down
cell walls and nuclear membranes to release the DNA
They work by chemically poking holes in the cell membranes or walls
Once holes are poked in the membranes the membranes can be further disrupted mechanically as with a blender
After that it is easier to get the contents of the cell out including the DNA
LETS HAVE A LOOK AT THE CELL
33
CELL TO DNA
httplearngeneticsutaheducontentlabsextractionhowtoHOW TO EXTRACT DNA FROM ANYTHING LIVING
34
THE WHYrsquoS WHY DETERGENT
Each cell is surrounded by a sack (the cell membrane)
DNA is found inside the second sack (nucleus) within each cell
To see the DNA we have to break it open A cell membrane has 2 layers of lipid(fat)
molecules with protein going through them When the lysis buffer (detergent) comes
close to the cell it captures the lipids and the proteins - breaks open the cell destroying the fatty membrane - DNA now released into the solution
35
A CELLS MEMBRANES HAVE TWO LAYERS OF LIPID (FAT) MOLECULES WITH PROTEINS GOING THROUGH THEM
36
Why detergent How does detergent work
Think about why you use soap to wash dishes or your hands To remove grease and dirt right
Soap molecules and grease molecules are made of two parts
1(Blue) Heads which like water 2(Green) Tails which hate water
37
AFTER ADDING THE DETERGENT WHAT DO YOU HAVE IN YOUR SOUP
38
THE WHYrsquoS ENZYME (MEAT TENDERIZER)
The DNA in the nucleus of the cell is molded folded and protected by proteins The tenderizer cut the proteins away from the DNA
In this experiment meat tenderizer acts as an enzyme to cut proteins just like a pair of scissors
The meat tenderizer cuts the proteins away from the DNA
39
THE WHYrsquoS CUTTING PROTEIN AND DNA
The DNA in the nucleus of the cell is moulded folded and protected by proteins
40
THE WHYrsquoSALCOHOL Alcohol is less dense than water so it floats
on top Look for clumps of white stringy stuff where the water and alcohol layers meet
DNA is not soluble in alcohol - other cell parts are
By adding alcohol DNA precipitates out of the solution and collect at the interface of the alcohol and soap layer
The colder the alcohol the less soluble the DNA will be in it
41
COLD ALCOHOL
DNA dissolves in water but precipitates in alcohol
Cold alcohol is used to separate DNA out of water-based solutions
This allows the DNA to be purified for subsequent genetic testing
Adding alcohol to a solution containing DNA is a simple way to obtain the pure DNA required and colder temperatures slow down enzymes that can break down DNA giving better extraction results
42
WHY IS DNA EXTRACTION IMPORTANT DNA extraction is an important molecular
biology procedure
By definition extraction is taking DNA out of any type of cell for the purpose of analysis
See Handouts for more information on the use of DNA extracted (extension only)
43
SUMMARIZEDNA EXTRACTING FROM KIWI FRUIT
44
BREAK DOWN CELL WALLS
Cells walls have to be destroyed to reach the DNA within cells
Extraction procedures to obtain pure DNA have to get rid of all the molecular and chemical components of the tissue from which the DNA is being extracted
First steps involve lysing or destroying the cell walls This can be done with a variety of chemical agents that are caustic to cell membranes but do not harm the DNA Cells can also be sonicated homogenized or ground up to destroy membranes
45
REMOVING Removing Lipids
Once the cell walls have been destroyed a detergent is added to get rid of the fats and oils that make up the cell membranes Detergents cause the fats and oils to dissolve into the solution
Remove ProteinsProteins and enzymes can be digested by
adding a protease to the solution Proteases break down proteins into small peptides and amino acids
46
PRECIPITATING AND PURIFYING Precipitate DNA
Adding cold alcohol to a solution will cause DNA to precipitate and aggregate The DNA can be collected by centrifuging the sample and pouring off the liquid layer The DNA should exist as a small pellet in the bottom of the centrifuge tube
Purify DNAThe DNA can be washed by re-suspending it
in a cold alcohol solution and re-centrifuging it several times to obtain a very pure DNA sample Typical alcohols used to precipitate DNA include ethanol and Isopropanol This process leaves a very pure sample for stringent DNA testing
47
PART B OBSERVING EXTRACTED DNA UNDER THE MICROSCOPE
48
PART B A CLOSER LOOK
8 Use a dropping pipette to carefully remove some of the threadlike substance from the top of your preparation
9 Place one or two drops onto the middle of a
microscope slide
10 Add two drops of methylene blue Wait 3 or 4 minutes to allow the methylene blue to be absorbed by the DNA
11 Carefully place a cover slip on the slide Gently press
a folded piece of paper towelling over the top of the prepared slide to soak up any excess liquid
12 Observe the DNA under low power then high power
49
DISCUSSIONANSWER IN GROUP RELATED WORKED 1 Write a detailed description of the material
floating at the top of the test tube after the alcohol was added
2 Describe the DNA as it appears under the
microscope under high power 3 Prepare a diagram of your DNA specimen 4 What was the reason for using methylene
blue
50
HOMEWORKCONSTRUCT A FLOW CHART DRAWING MIND MAP OF THE PROCESS YOU USED EXTRACTING THE DNA FROM KIWIrsquoS
Next to each step explain how the method you used was important (Hint What substances make up the membranes of cells and cell organelles
How do detergents work
What is the active ingredient in meat tenderiser
51
TIPS STEPS FOR FLOW CHART In order to release the DNA from the
nuclei of the pea cells you first separated the cells from one another
Then the cell and nuclear membranes needed to be ruptured to release the cell contents and the contents of the nucleus
Once removed from the nuclear membrane the DNA had to be untangled into the visible threadlike structures you ended up with
52
HOMEWORK EXTENSION GROUP QUESTIONS Where can DNA be found in the cell Discuss the action of the soap (detergent) on the cell
What is the purpose of the soap in this activity What was the purpose of the Sodium Chloride Include a
discussion of polarity and charged particles Why was the cold ethanol added to the soap and salt
mixture Describe the appearance of your final product Draw a diagram of DNA containing 5 sets of nucleotide
bases labelling the hydrogen bonds between the bases References and Resources Adapted from Berry Full
of DNA by Diane Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
53
HTTPWWWGENOMEBCCAEDUCATIONTEACHERSCLASSROOM-ACTIVITIESDNA-EXTRACTION
54
EXTRACHROMOSOMES EXIST IN PAIRS Because our chromosomes exist in pairs (and consequently we
have 2 alleles of each gene) we are a diploid species This is why our somatic cells are represented as 2n Our gametes (sperm and ova) on the other hand are haploid and are represented as n Other species may have different ploidy for example
triploid (3n) seedless watermelons tetaploid (4n) salmonidae fish pentaploid (5n) Kenai birch hexaploid (6n) some types of wheat kiwi fruit octaploid (8n) acipenser (a genus
of sturgeon fish strawberies) decaploid (10n) some strawberries dodecaploid (12n) some types of amphibians eg Xenopus
ruwenzoriensis
wwwcyberusca
55
YOUR EXTRACTED DNA UNDER THE MICROSCOPE LOOKED
LIKE
Collaboration-Brainstorming on what to look at under the
- DNA extraction- DNA is too small for even a microscope to see and after the extraction it just appears like a blob True or false
Could you any DNA strand
wwwemployeescsbsjuedu
56
HANDOUT ON DNA EXTRACTIONBACKGROUND EVERY DAY LIFE
EXTRA READING MATERIAL(EXTENSION )
DISCOVERING DNA DNA ON THE INSIDE USE OF DNA EXTRACTION INN EVERYDAY LIFE WHY IS DNA TESTING GOOD MOLECULAR BIOLOGY PCR-based diagnostics of genomic DNA
57
REFERENCES httpwwwsquidoocomhow-to-get-dna-from-a-kiwi-fruit Why Is DNA Testing Good | eHowcom
httpwwwehowcomabout_6684410_dna-testing-good_htmlixzz1MkbcIJs5 Why Is DNA Extraction Important | eHowcom
httpwwwehowcomlist_5839095_dna-extraction-important_htmlixzz1MkZDrqyY
Why Is Sodium Used in DNA Extraction | eHowcom httpwwwehowcomabout_6504902_sodium-used-dna-extraction_htmlixzz1MkaGWg57
Why Is Cold Alcohol Used in DNA Tests | eHowcom httpwwwehowcomabout_6399349_cold-alcohol-used-dna-tests_htmlixzz1MkbEmEdu
httplearngeneticsutaheducontentlabsextractionhowto www kamibudaksainsblogspotcom
httpwwwchemwatch goldcom References and Resources Adapted from Berry Full of DNA by Diane
Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
58
REFERENCES
Uses of DNA Extraction | eHowcom httpwwwehowcomabout_5344428_uses-dna-extractionhtmlixzz1MkWQB2TZ
Science 10 A Contextual Approach Heineman Queensland Science Projects Regan Spence Maggie Spenceley
httpenwikipediaorgwikiMolecular_biology httpwwwclinchemorgcgicontent
extract44102201 http
employeescsbsjueduhjakubowskiclassesch331dnaoldnastructurehtml
httpwwwgenomebccaeducationteachersclassroom-activitiesdna-extraction
18
PART A EXTRACTING THE DNA
METHOD
1 Peal and Place the kiwi and water in the beaker and mush it with a mortar
and pestle DNA SOURCE About 125 ml 12 cup Twice as much water as DNA source (250
ml 1 cup) Table salt large pinch 1g 14 teaspoon Stir until the mixture is of a thin soupy
consistency
TAKE NOTES ndash NO TEXTBOOKS ALLOWED WHEN PERFORMING EXPERIMENT ndashMUST RECALL PROCEDURES METHOD
19
WHAT MUST I DO NOW
STRAIN THE DNA MIXTURE
DISHWASHING DETERGENT
20
2 Pour the thin cell mixture through the kitchen strainer (cheesecloth) into another large beaker
3 Measure 12 ml of the soup into
a small beakerAdd 2 ml of liquid detergentSwirl to mix stir thoroughly using a
glass rodLet mixture sit in container with
hot tap water (60 - 65degC) for 5 - 10 minutes
21
WHAT MUST I DO NOWMEAT TENDERIZER
ENZYME POWDERALCOHOL SEPARATION
wwwforumsoverclockerscomau
22
4 Add a pinch of enzyme (meat tenderizer) to mixture
(Using pineapple juice or contact lens cleaning solution will do the same as tenderizer)
Gently stir with toothpick skewer for 5 minutes continue stirring not too vigorously Be careful If you stir too hard youll break up the DNA making it harder to see
5 Quarter-fill a large test tube with the mixture
23
SLOWLY pour the same amount ice - cold (70 - 95 isopropyl or ethyl alcohol) into test tube pour alcohol down the side of the test tube
Tilting the test tube will make this easier to do
It forms a layer on top of the cell mixture
DO NOT MIX THE TWO LAYERS TOGETHER
Amount of alcohol and mixture should be the same
24
SEPARATION USING ALCOHOL
25
7 Observe the mixture for a few minutes You will see a white threadlike substance rise from the mixture to rest above in the alcohol layer
This is the DNA that you have extracted from the cells of the kiwi fruit
8 You can get more DNA to precipitate from the solution using a DNA collecting tool (glass or paper clip hook or cut inoculation needle)
Gently lift the water solution up into the alcohol layer (this allows more DNA to get in contact with the alcohol and precipitate)
26
INFORMATION DNA precipitates as a white stringy
ldquosnottyrdquo film at the water - alcohol interface and eventually will rise into the alcohol layer from the mixture layer
Allow test tube to sit for several minutes The clearer the DNA is the fewer
impurities you have
If you have an acceptable amount of DNA it can be spooled by rotating your collecting tool and then transferred into a clean tube container
27
WOW ndash SPOOLING THE DNA If you are careful you may be able wind up the DNA
around a glass rod or a skewer Position the tip of the glass rod or skewer where you can see the threads of DNA Steadily twist the rod or skewer as if you were making candy floss Alternatively use a straw to pull it out by suction ndash be careful not to get in you mouth
Donrsquot go too quickly
You should be able to pull the strands of DNA out of the mixture
ASK STUDENTS TO TAKE MOBILE PHONE PICTURES OF THEIR OWN EXTRACTED DNA AND COMPARE APPEARANCE WITH OTHERS
Photos of steps can be inserted in flow diagram(ICT)
28
KIWIDNA
29
If you want to save your DNA you can transfer it to a small container filled with alcohol
Leave tube container uncapped until the ethanol has evaporated
DNA can be stored in the fridge (dry or water buffer can be added)
You can now investigate the property
30
WHAT IS THAT STRINGY STUFFDNA IS A LONG STRINGY MOLECULE THE SALT THAT YOU ADDED IN STEP ONE HELPS IT STICK TOGETHER SO WHAT YOU SEE ARE CLUMPS OF TANGLED DNA MOLECULES
DNA NORMALLY STAYS DISSOLVED IN WATER BUT WHEN SALTY DNA COMES IN CONTACT WITH ALCOHOL IT BECOMES UNDISSOLVED THIS IS CALLED PRECIPITATION THE PHYSICAL FORCE OF THE DNA CLUMPING TOGETHER AS IT PRECIPITATES PULLS MORE STRANDS ALONG WITH IT AS IT RISES INTO THE ALCOHOL
31
THE WHYrsquoS Blending separated the pea cells In order to extract DNA from a cell the associated
membranes and proteins must first be removed (break apart the cells) and then physically separated (loosen the tough cell wall) from the DNA
To see the DNA we have to break open these two sacks We do this with detergent and salt( Sodium can be
involved in several of the steps ) Sodium is an element Its chemical symbol is Na for
Natrium the Latin word for sodium It is a positive ion and often associates with negative ions as part of useful compounds
Salt help precipitate protein and carbohydrates away from the DNA
Salt helps strip away the proteins associated with DNA + and ndash charge
32
THE WHYrsquoS Salt and Detergents are used to break down
cell walls and nuclear membranes to release the DNA
They work by chemically poking holes in the cell membranes or walls
Once holes are poked in the membranes the membranes can be further disrupted mechanically as with a blender
After that it is easier to get the contents of the cell out including the DNA
LETS HAVE A LOOK AT THE CELL
33
CELL TO DNA
httplearngeneticsutaheducontentlabsextractionhowtoHOW TO EXTRACT DNA FROM ANYTHING LIVING
34
THE WHYrsquoS WHY DETERGENT
Each cell is surrounded by a sack (the cell membrane)
DNA is found inside the second sack (nucleus) within each cell
To see the DNA we have to break it open A cell membrane has 2 layers of lipid(fat)
molecules with protein going through them When the lysis buffer (detergent) comes
close to the cell it captures the lipids and the proteins - breaks open the cell destroying the fatty membrane - DNA now released into the solution
35
A CELLS MEMBRANES HAVE TWO LAYERS OF LIPID (FAT) MOLECULES WITH PROTEINS GOING THROUGH THEM
36
Why detergent How does detergent work
Think about why you use soap to wash dishes or your hands To remove grease and dirt right
Soap molecules and grease molecules are made of two parts
1(Blue) Heads which like water 2(Green) Tails which hate water
37
AFTER ADDING THE DETERGENT WHAT DO YOU HAVE IN YOUR SOUP
38
THE WHYrsquoS ENZYME (MEAT TENDERIZER)
The DNA in the nucleus of the cell is molded folded and protected by proteins The tenderizer cut the proteins away from the DNA
In this experiment meat tenderizer acts as an enzyme to cut proteins just like a pair of scissors
The meat tenderizer cuts the proteins away from the DNA
39
THE WHYrsquoS CUTTING PROTEIN AND DNA
The DNA in the nucleus of the cell is moulded folded and protected by proteins
40
THE WHYrsquoSALCOHOL Alcohol is less dense than water so it floats
on top Look for clumps of white stringy stuff where the water and alcohol layers meet
DNA is not soluble in alcohol - other cell parts are
By adding alcohol DNA precipitates out of the solution and collect at the interface of the alcohol and soap layer
The colder the alcohol the less soluble the DNA will be in it
41
COLD ALCOHOL
DNA dissolves in water but precipitates in alcohol
Cold alcohol is used to separate DNA out of water-based solutions
This allows the DNA to be purified for subsequent genetic testing
Adding alcohol to a solution containing DNA is a simple way to obtain the pure DNA required and colder temperatures slow down enzymes that can break down DNA giving better extraction results
42
WHY IS DNA EXTRACTION IMPORTANT DNA extraction is an important molecular
biology procedure
By definition extraction is taking DNA out of any type of cell for the purpose of analysis
See Handouts for more information on the use of DNA extracted (extension only)
43
SUMMARIZEDNA EXTRACTING FROM KIWI FRUIT
44
BREAK DOWN CELL WALLS
Cells walls have to be destroyed to reach the DNA within cells
Extraction procedures to obtain pure DNA have to get rid of all the molecular and chemical components of the tissue from which the DNA is being extracted
First steps involve lysing or destroying the cell walls This can be done with a variety of chemical agents that are caustic to cell membranes but do not harm the DNA Cells can also be sonicated homogenized or ground up to destroy membranes
45
REMOVING Removing Lipids
Once the cell walls have been destroyed a detergent is added to get rid of the fats and oils that make up the cell membranes Detergents cause the fats and oils to dissolve into the solution
Remove ProteinsProteins and enzymes can be digested by
adding a protease to the solution Proteases break down proteins into small peptides and amino acids
46
PRECIPITATING AND PURIFYING Precipitate DNA
Adding cold alcohol to a solution will cause DNA to precipitate and aggregate The DNA can be collected by centrifuging the sample and pouring off the liquid layer The DNA should exist as a small pellet in the bottom of the centrifuge tube
Purify DNAThe DNA can be washed by re-suspending it
in a cold alcohol solution and re-centrifuging it several times to obtain a very pure DNA sample Typical alcohols used to precipitate DNA include ethanol and Isopropanol This process leaves a very pure sample for stringent DNA testing
47
PART B OBSERVING EXTRACTED DNA UNDER THE MICROSCOPE
48
PART B A CLOSER LOOK
8 Use a dropping pipette to carefully remove some of the threadlike substance from the top of your preparation
9 Place one or two drops onto the middle of a
microscope slide
10 Add two drops of methylene blue Wait 3 or 4 minutes to allow the methylene blue to be absorbed by the DNA
11 Carefully place a cover slip on the slide Gently press
a folded piece of paper towelling over the top of the prepared slide to soak up any excess liquid
12 Observe the DNA under low power then high power
49
DISCUSSIONANSWER IN GROUP RELATED WORKED 1 Write a detailed description of the material
floating at the top of the test tube after the alcohol was added
2 Describe the DNA as it appears under the
microscope under high power 3 Prepare a diagram of your DNA specimen 4 What was the reason for using methylene
blue
50
HOMEWORKCONSTRUCT A FLOW CHART DRAWING MIND MAP OF THE PROCESS YOU USED EXTRACTING THE DNA FROM KIWIrsquoS
Next to each step explain how the method you used was important (Hint What substances make up the membranes of cells and cell organelles
How do detergents work
What is the active ingredient in meat tenderiser
51
TIPS STEPS FOR FLOW CHART In order to release the DNA from the
nuclei of the pea cells you first separated the cells from one another
Then the cell and nuclear membranes needed to be ruptured to release the cell contents and the contents of the nucleus
Once removed from the nuclear membrane the DNA had to be untangled into the visible threadlike structures you ended up with
52
HOMEWORK EXTENSION GROUP QUESTIONS Where can DNA be found in the cell Discuss the action of the soap (detergent) on the cell
What is the purpose of the soap in this activity What was the purpose of the Sodium Chloride Include a
discussion of polarity and charged particles Why was the cold ethanol added to the soap and salt
mixture Describe the appearance of your final product Draw a diagram of DNA containing 5 sets of nucleotide
bases labelling the hydrogen bonds between the bases References and Resources Adapted from Berry Full
of DNA by Diane Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
53
HTTPWWWGENOMEBCCAEDUCATIONTEACHERSCLASSROOM-ACTIVITIESDNA-EXTRACTION
54
EXTRACHROMOSOMES EXIST IN PAIRS Because our chromosomes exist in pairs (and consequently we
have 2 alleles of each gene) we are a diploid species This is why our somatic cells are represented as 2n Our gametes (sperm and ova) on the other hand are haploid and are represented as n Other species may have different ploidy for example
triploid (3n) seedless watermelons tetaploid (4n) salmonidae fish pentaploid (5n) Kenai birch hexaploid (6n) some types of wheat kiwi fruit octaploid (8n) acipenser (a genus
of sturgeon fish strawberies) decaploid (10n) some strawberries dodecaploid (12n) some types of amphibians eg Xenopus
ruwenzoriensis
wwwcyberusca
55
YOUR EXTRACTED DNA UNDER THE MICROSCOPE LOOKED
LIKE
Collaboration-Brainstorming on what to look at under the
- DNA extraction- DNA is too small for even a microscope to see and after the extraction it just appears like a blob True or false
Could you any DNA strand
wwwemployeescsbsjuedu
56
HANDOUT ON DNA EXTRACTIONBACKGROUND EVERY DAY LIFE
EXTRA READING MATERIAL(EXTENSION )
DISCOVERING DNA DNA ON THE INSIDE USE OF DNA EXTRACTION INN EVERYDAY LIFE WHY IS DNA TESTING GOOD MOLECULAR BIOLOGY PCR-based diagnostics of genomic DNA
57
REFERENCES httpwwwsquidoocomhow-to-get-dna-from-a-kiwi-fruit Why Is DNA Testing Good | eHowcom
httpwwwehowcomabout_6684410_dna-testing-good_htmlixzz1MkbcIJs5 Why Is DNA Extraction Important | eHowcom
httpwwwehowcomlist_5839095_dna-extraction-important_htmlixzz1MkZDrqyY
Why Is Sodium Used in DNA Extraction | eHowcom httpwwwehowcomabout_6504902_sodium-used-dna-extraction_htmlixzz1MkaGWg57
Why Is Cold Alcohol Used in DNA Tests | eHowcom httpwwwehowcomabout_6399349_cold-alcohol-used-dna-tests_htmlixzz1MkbEmEdu
httplearngeneticsutaheducontentlabsextractionhowto www kamibudaksainsblogspotcom
httpwwwchemwatch goldcom References and Resources Adapted from Berry Full of DNA by Diane
Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
58
REFERENCES
Uses of DNA Extraction | eHowcom httpwwwehowcomabout_5344428_uses-dna-extractionhtmlixzz1MkWQB2TZ
Science 10 A Contextual Approach Heineman Queensland Science Projects Regan Spence Maggie Spenceley
httpenwikipediaorgwikiMolecular_biology httpwwwclinchemorgcgicontent
extract44102201 http
employeescsbsjueduhjakubowskiclassesch331dnaoldnastructurehtml
httpwwwgenomebccaeducationteachersclassroom-activitiesdna-extraction
19
WHAT MUST I DO NOW
STRAIN THE DNA MIXTURE
DISHWASHING DETERGENT
20
2 Pour the thin cell mixture through the kitchen strainer (cheesecloth) into another large beaker
3 Measure 12 ml of the soup into
a small beakerAdd 2 ml of liquid detergentSwirl to mix stir thoroughly using a
glass rodLet mixture sit in container with
hot tap water (60 - 65degC) for 5 - 10 minutes
21
WHAT MUST I DO NOWMEAT TENDERIZER
ENZYME POWDERALCOHOL SEPARATION
wwwforumsoverclockerscomau
22
4 Add a pinch of enzyme (meat tenderizer) to mixture
(Using pineapple juice or contact lens cleaning solution will do the same as tenderizer)
Gently stir with toothpick skewer for 5 minutes continue stirring not too vigorously Be careful If you stir too hard youll break up the DNA making it harder to see
5 Quarter-fill a large test tube with the mixture
23
SLOWLY pour the same amount ice - cold (70 - 95 isopropyl or ethyl alcohol) into test tube pour alcohol down the side of the test tube
Tilting the test tube will make this easier to do
It forms a layer on top of the cell mixture
DO NOT MIX THE TWO LAYERS TOGETHER
Amount of alcohol and mixture should be the same
24
SEPARATION USING ALCOHOL
25
7 Observe the mixture for a few minutes You will see a white threadlike substance rise from the mixture to rest above in the alcohol layer
This is the DNA that you have extracted from the cells of the kiwi fruit
8 You can get more DNA to precipitate from the solution using a DNA collecting tool (glass or paper clip hook or cut inoculation needle)
Gently lift the water solution up into the alcohol layer (this allows more DNA to get in contact with the alcohol and precipitate)
26
INFORMATION DNA precipitates as a white stringy
ldquosnottyrdquo film at the water - alcohol interface and eventually will rise into the alcohol layer from the mixture layer
Allow test tube to sit for several minutes The clearer the DNA is the fewer
impurities you have
If you have an acceptable amount of DNA it can be spooled by rotating your collecting tool and then transferred into a clean tube container
27
WOW ndash SPOOLING THE DNA If you are careful you may be able wind up the DNA
around a glass rod or a skewer Position the tip of the glass rod or skewer where you can see the threads of DNA Steadily twist the rod or skewer as if you were making candy floss Alternatively use a straw to pull it out by suction ndash be careful not to get in you mouth
Donrsquot go too quickly
You should be able to pull the strands of DNA out of the mixture
ASK STUDENTS TO TAKE MOBILE PHONE PICTURES OF THEIR OWN EXTRACTED DNA AND COMPARE APPEARANCE WITH OTHERS
Photos of steps can be inserted in flow diagram(ICT)
28
KIWIDNA
29
If you want to save your DNA you can transfer it to a small container filled with alcohol
Leave tube container uncapped until the ethanol has evaporated
DNA can be stored in the fridge (dry or water buffer can be added)
You can now investigate the property
30
WHAT IS THAT STRINGY STUFFDNA IS A LONG STRINGY MOLECULE THE SALT THAT YOU ADDED IN STEP ONE HELPS IT STICK TOGETHER SO WHAT YOU SEE ARE CLUMPS OF TANGLED DNA MOLECULES
DNA NORMALLY STAYS DISSOLVED IN WATER BUT WHEN SALTY DNA COMES IN CONTACT WITH ALCOHOL IT BECOMES UNDISSOLVED THIS IS CALLED PRECIPITATION THE PHYSICAL FORCE OF THE DNA CLUMPING TOGETHER AS IT PRECIPITATES PULLS MORE STRANDS ALONG WITH IT AS IT RISES INTO THE ALCOHOL
31
THE WHYrsquoS Blending separated the pea cells In order to extract DNA from a cell the associated
membranes and proteins must first be removed (break apart the cells) and then physically separated (loosen the tough cell wall) from the DNA
To see the DNA we have to break open these two sacks We do this with detergent and salt( Sodium can be
involved in several of the steps ) Sodium is an element Its chemical symbol is Na for
Natrium the Latin word for sodium It is a positive ion and often associates with negative ions as part of useful compounds
Salt help precipitate protein and carbohydrates away from the DNA
Salt helps strip away the proteins associated with DNA + and ndash charge
32
THE WHYrsquoS Salt and Detergents are used to break down
cell walls and nuclear membranes to release the DNA
They work by chemically poking holes in the cell membranes or walls
Once holes are poked in the membranes the membranes can be further disrupted mechanically as with a blender
After that it is easier to get the contents of the cell out including the DNA
LETS HAVE A LOOK AT THE CELL
33
CELL TO DNA
httplearngeneticsutaheducontentlabsextractionhowtoHOW TO EXTRACT DNA FROM ANYTHING LIVING
34
THE WHYrsquoS WHY DETERGENT
Each cell is surrounded by a sack (the cell membrane)
DNA is found inside the second sack (nucleus) within each cell
To see the DNA we have to break it open A cell membrane has 2 layers of lipid(fat)
molecules with protein going through them When the lysis buffer (detergent) comes
close to the cell it captures the lipids and the proteins - breaks open the cell destroying the fatty membrane - DNA now released into the solution
35
A CELLS MEMBRANES HAVE TWO LAYERS OF LIPID (FAT) MOLECULES WITH PROTEINS GOING THROUGH THEM
36
Why detergent How does detergent work
Think about why you use soap to wash dishes or your hands To remove grease and dirt right
Soap molecules and grease molecules are made of two parts
1(Blue) Heads which like water 2(Green) Tails which hate water
37
AFTER ADDING THE DETERGENT WHAT DO YOU HAVE IN YOUR SOUP
38
THE WHYrsquoS ENZYME (MEAT TENDERIZER)
The DNA in the nucleus of the cell is molded folded and protected by proteins The tenderizer cut the proteins away from the DNA
In this experiment meat tenderizer acts as an enzyme to cut proteins just like a pair of scissors
The meat tenderizer cuts the proteins away from the DNA
39
THE WHYrsquoS CUTTING PROTEIN AND DNA
The DNA in the nucleus of the cell is moulded folded and protected by proteins
40
THE WHYrsquoSALCOHOL Alcohol is less dense than water so it floats
on top Look for clumps of white stringy stuff where the water and alcohol layers meet
DNA is not soluble in alcohol - other cell parts are
By adding alcohol DNA precipitates out of the solution and collect at the interface of the alcohol and soap layer
The colder the alcohol the less soluble the DNA will be in it
41
COLD ALCOHOL
DNA dissolves in water but precipitates in alcohol
Cold alcohol is used to separate DNA out of water-based solutions
This allows the DNA to be purified for subsequent genetic testing
Adding alcohol to a solution containing DNA is a simple way to obtain the pure DNA required and colder temperatures slow down enzymes that can break down DNA giving better extraction results
42
WHY IS DNA EXTRACTION IMPORTANT DNA extraction is an important molecular
biology procedure
By definition extraction is taking DNA out of any type of cell for the purpose of analysis
See Handouts for more information on the use of DNA extracted (extension only)
43
SUMMARIZEDNA EXTRACTING FROM KIWI FRUIT
44
BREAK DOWN CELL WALLS
Cells walls have to be destroyed to reach the DNA within cells
Extraction procedures to obtain pure DNA have to get rid of all the molecular and chemical components of the tissue from which the DNA is being extracted
First steps involve lysing or destroying the cell walls This can be done with a variety of chemical agents that are caustic to cell membranes but do not harm the DNA Cells can also be sonicated homogenized or ground up to destroy membranes
45
REMOVING Removing Lipids
Once the cell walls have been destroyed a detergent is added to get rid of the fats and oils that make up the cell membranes Detergents cause the fats and oils to dissolve into the solution
Remove ProteinsProteins and enzymes can be digested by
adding a protease to the solution Proteases break down proteins into small peptides and amino acids
46
PRECIPITATING AND PURIFYING Precipitate DNA
Adding cold alcohol to a solution will cause DNA to precipitate and aggregate The DNA can be collected by centrifuging the sample and pouring off the liquid layer The DNA should exist as a small pellet in the bottom of the centrifuge tube
Purify DNAThe DNA can be washed by re-suspending it
in a cold alcohol solution and re-centrifuging it several times to obtain a very pure DNA sample Typical alcohols used to precipitate DNA include ethanol and Isopropanol This process leaves a very pure sample for stringent DNA testing
47
PART B OBSERVING EXTRACTED DNA UNDER THE MICROSCOPE
48
PART B A CLOSER LOOK
8 Use a dropping pipette to carefully remove some of the threadlike substance from the top of your preparation
9 Place one or two drops onto the middle of a
microscope slide
10 Add two drops of methylene blue Wait 3 or 4 minutes to allow the methylene blue to be absorbed by the DNA
11 Carefully place a cover slip on the slide Gently press
a folded piece of paper towelling over the top of the prepared slide to soak up any excess liquid
12 Observe the DNA under low power then high power
49
DISCUSSIONANSWER IN GROUP RELATED WORKED 1 Write a detailed description of the material
floating at the top of the test tube after the alcohol was added
2 Describe the DNA as it appears under the
microscope under high power 3 Prepare a diagram of your DNA specimen 4 What was the reason for using methylene
blue
50
HOMEWORKCONSTRUCT A FLOW CHART DRAWING MIND MAP OF THE PROCESS YOU USED EXTRACTING THE DNA FROM KIWIrsquoS
Next to each step explain how the method you used was important (Hint What substances make up the membranes of cells and cell organelles
How do detergents work
What is the active ingredient in meat tenderiser
51
TIPS STEPS FOR FLOW CHART In order to release the DNA from the
nuclei of the pea cells you first separated the cells from one another
Then the cell and nuclear membranes needed to be ruptured to release the cell contents and the contents of the nucleus
Once removed from the nuclear membrane the DNA had to be untangled into the visible threadlike structures you ended up with
52
HOMEWORK EXTENSION GROUP QUESTIONS Where can DNA be found in the cell Discuss the action of the soap (detergent) on the cell
What is the purpose of the soap in this activity What was the purpose of the Sodium Chloride Include a
discussion of polarity and charged particles Why was the cold ethanol added to the soap and salt
mixture Describe the appearance of your final product Draw a diagram of DNA containing 5 sets of nucleotide
bases labelling the hydrogen bonds between the bases References and Resources Adapted from Berry Full
of DNA by Diane Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
53
HTTPWWWGENOMEBCCAEDUCATIONTEACHERSCLASSROOM-ACTIVITIESDNA-EXTRACTION
54
EXTRACHROMOSOMES EXIST IN PAIRS Because our chromosomes exist in pairs (and consequently we
have 2 alleles of each gene) we are a diploid species This is why our somatic cells are represented as 2n Our gametes (sperm and ova) on the other hand are haploid and are represented as n Other species may have different ploidy for example
triploid (3n) seedless watermelons tetaploid (4n) salmonidae fish pentaploid (5n) Kenai birch hexaploid (6n) some types of wheat kiwi fruit octaploid (8n) acipenser (a genus
of sturgeon fish strawberies) decaploid (10n) some strawberries dodecaploid (12n) some types of amphibians eg Xenopus
ruwenzoriensis
wwwcyberusca
55
YOUR EXTRACTED DNA UNDER THE MICROSCOPE LOOKED
LIKE
Collaboration-Brainstorming on what to look at under the
- DNA extraction- DNA is too small for even a microscope to see and after the extraction it just appears like a blob True or false
Could you any DNA strand
wwwemployeescsbsjuedu
56
HANDOUT ON DNA EXTRACTIONBACKGROUND EVERY DAY LIFE
EXTRA READING MATERIAL(EXTENSION )
DISCOVERING DNA DNA ON THE INSIDE USE OF DNA EXTRACTION INN EVERYDAY LIFE WHY IS DNA TESTING GOOD MOLECULAR BIOLOGY PCR-based diagnostics of genomic DNA
57
REFERENCES httpwwwsquidoocomhow-to-get-dna-from-a-kiwi-fruit Why Is DNA Testing Good | eHowcom
httpwwwehowcomabout_6684410_dna-testing-good_htmlixzz1MkbcIJs5 Why Is DNA Extraction Important | eHowcom
httpwwwehowcomlist_5839095_dna-extraction-important_htmlixzz1MkZDrqyY
Why Is Sodium Used in DNA Extraction | eHowcom httpwwwehowcomabout_6504902_sodium-used-dna-extraction_htmlixzz1MkaGWg57
Why Is Cold Alcohol Used in DNA Tests | eHowcom httpwwwehowcomabout_6399349_cold-alcohol-used-dna-tests_htmlixzz1MkbEmEdu
httplearngeneticsutaheducontentlabsextractionhowto www kamibudaksainsblogspotcom
httpwwwchemwatch goldcom References and Resources Adapted from Berry Full of DNA by Diane
Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
58
REFERENCES
Uses of DNA Extraction | eHowcom httpwwwehowcomabout_5344428_uses-dna-extractionhtmlixzz1MkWQB2TZ
Science 10 A Contextual Approach Heineman Queensland Science Projects Regan Spence Maggie Spenceley
httpenwikipediaorgwikiMolecular_biology httpwwwclinchemorgcgicontent
extract44102201 http
employeescsbsjueduhjakubowskiclassesch331dnaoldnastructurehtml
httpwwwgenomebccaeducationteachersclassroom-activitiesdna-extraction
20
2 Pour the thin cell mixture through the kitchen strainer (cheesecloth) into another large beaker
3 Measure 12 ml of the soup into
a small beakerAdd 2 ml of liquid detergentSwirl to mix stir thoroughly using a
glass rodLet mixture sit in container with
hot tap water (60 - 65degC) for 5 - 10 minutes
21
WHAT MUST I DO NOWMEAT TENDERIZER
ENZYME POWDERALCOHOL SEPARATION
wwwforumsoverclockerscomau
22
4 Add a pinch of enzyme (meat tenderizer) to mixture
(Using pineapple juice or contact lens cleaning solution will do the same as tenderizer)
Gently stir with toothpick skewer for 5 minutes continue stirring not too vigorously Be careful If you stir too hard youll break up the DNA making it harder to see
5 Quarter-fill a large test tube with the mixture
23
SLOWLY pour the same amount ice - cold (70 - 95 isopropyl or ethyl alcohol) into test tube pour alcohol down the side of the test tube
Tilting the test tube will make this easier to do
It forms a layer on top of the cell mixture
DO NOT MIX THE TWO LAYERS TOGETHER
Amount of alcohol and mixture should be the same
24
SEPARATION USING ALCOHOL
25
7 Observe the mixture for a few minutes You will see a white threadlike substance rise from the mixture to rest above in the alcohol layer
This is the DNA that you have extracted from the cells of the kiwi fruit
8 You can get more DNA to precipitate from the solution using a DNA collecting tool (glass or paper clip hook or cut inoculation needle)
Gently lift the water solution up into the alcohol layer (this allows more DNA to get in contact with the alcohol and precipitate)
26
INFORMATION DNA precipitates as a white stringy
ldquosnottyrdquo film at the water - alcohol interface and eventually will rise into the alcohol layer from the mixture layer
Allow test tube to sit for several minutes The clearer the DNA is the fewer
impurities you have
If you have an acceptable amount of DNA it can be spooled by rotating your collecting tool and then transferred into a clean tube container
27
WOW ndash SPOOLING THE DNA If you are careful you may be able wind up the DNA
around a glass rod or a skewer Position the tip of the glass rod or skewer where you can see the threads of DNA Steadily twist the rod or skewer as if you were making candy floss Alternatively use a straw to pull it out by suction ndash be careful not to get in you mouth
Donrsquot go too quickly
You should be able to pull the strands of DNA out of the mixture
ASK STUDENTS TO TAKE MOBILE PHONE PICTURES OF THEIR OWN EXTRACTED DNA AND COMPARE APPEARANCE WITH OTHERS
Photos of steps can be inserted in flow diagram(ICT)
28
KIWIDNA
29
If you want to save your DNA you can transfer it to a small container filled with alcohol
Leave tube container uncapped until the ethanol has evaporated
DNA can be stored in the fridge (dry or water buffer can be added)
You can now investigate the property
30
WHAT IS THAT STRINGY STUFFDNA IS A LONG STRINGY MOLECULE THE SALT THAT YOU ADDED IN STEP ONE HELPS IT STICK TOGETHER SO WHAT YOU SEE ARE CLUMPS OF TANGLED DNA MOLECULES
DNA NORMALLY STAYS DISSOLVED IN WATER BUT WHEN SALTY DNA COMES IN CONTACT WITH ALCOHOL IT BECOMES UNDISSOLVED THIS IS CALLED PRECIPITATION THE PHYSICAL FORCE OF THE DNA CLUMPING TOGETHER AS IT PRECIPITATES PULLS MORE STRANDS ALONG WITH IT AS IT RISES INTO THE ALCOHOL
31
THE WHYrsquoS Blending separated the pea cells In order to extract DNA from a cell the associated
membranes and proteins must first be removed (break apart the cells) and then physically separated (loosen the tough cell wall) from the DNA
To see the DNA we have to break open these two sacks We do this with detergent and salt( Sodium can be
involved in several of the steps ) Sodium is an element Its chemical symbol is Na for
Natrium the Latin word for sodium It is a positive ion and often associates with negative ions as part of useful compounds
Salt help precipitate protein and carbohydrates away from the DNA
Salt helps strip away the proteins associated with DNA + and ndash charge
32
THE WHYrsquoS Salt and Detergents are used to break down
cell walls and nuclear membranes to release the DNA
They work by chemically poking holes in the cell membranes or walls
Once holes are poked in the membranes the membranes can be further disrupted mechanically as with a blender
After that it is easier to get the contents of the cell out including the DNA
LETS HAVE A LOOK AT THE CELL
33
CELL TO DNA
httplearngeneticsutaheducontentlabsextractionhowtoHOW TO EXTRACT DNA FROM ANYTHING LIVING
34
THE WHYrsquoS WHY DETERGENT
Each cell is surrounded by a sack (the cell membrane)
DNA is found inside the second sack (nucleus) within each cell
To see the DNA we have to break it open A cell membrane has 2 layers of lipid(fat)
molecules with protein going through them When the lysis buffer (detergent) comes
close to the cell it captures the lipids and the proteins - breaks open the cell destroying the fatty membrane - DNA now released into the solution
35
A CELLS MEMBRANES HAVE TWO LAYERS OF LIPID (FAT) MOLECULES WITH PROTEINS GOING THROUGH THEM
36
Why detergent How does detergent work
Think about why you use soap to wash dishes or your hands To remove grease and dirt right
Soap molecules and grease molecules are made of two parts
1(Blue) Heads which like water 2(Green) Tails which hate water
37
AFTER ADDING THE DETERGENT WHAT DO YOU HAVE IN YOUR SOUP
38
THE WHYrsquoS ENZYME (MEAT TENDERIZER)
The DNA in the nucleus of the cell is molded folded and protected by proteins The tenderizer cut the proteins away from the DNA
In this experiment meat tenderizer acts as an enzyme to cut proteins just like a pair of scissors
The meat tenderizer cuts the proteins away from the DNA
39
THE WHYrsquoS CUTTING PROTEIN AND DNA
The DNA in the nucleus of the cell is moulded folded and protected by proteins
40
THE WHYrsquoSALCOHOL Alcohol is less dense than water so it floats
on top Look for clumps of white stringy stuff where the water and alcohol layers meet
DNA is not soluble in alcohol - other cell parts are
By adding alcohol DNA precipitates out of the solution and collect at the interface of the alcohol and soap layer
The colder the alcohol the less soluble the DNA will be in it
41
COLD ALCOHOL
DNA dissolves in water but precipitates in alcohol
Cold alcohol is used to separate DNA out of water-based solutions
This allows the DNA to be purified for subsequent genetic testing
Adding alcohol to a solution containing DNA is a simple way to obtain the pure DNA required and colder temperatures slow down enzymes that can break down DNA giving better extraction results
42
WHY IS DNA EXTRACTION IMPORTANT DNA extraction is an important molecular
biology procedure
By definition extraction is taking DNA out of any type of cell for the purpose of analysis
See Handouts for more information on the use of DNA extracted (extension only)
43
SUMMARIZEDNA EXTRACTING FROM KIWI FRUIT
44
BREAK DOWN CELL WALLS
Cells walls have to be destroyed to reach the DNA within cells
Extraction procedures to obtain pure DNA have to get rid of all the molecular and chemical components of the tissue from which the DNA is being extracted
First steps involve lysing or destroying the cell walls This can be done with a variety of chemical agents that are caustic to cell membranes but do not harm the DNA Cells can also be sonicated homogenized or ground up to destroy membranes
45
REMOVING Removing Lipids
Once the cell walls have been destroyed a detergent is added to get rid of the fats and oils that make up the cell membranes Detergents cause the fats and oils to dissolve into the solution
Remove ProteinsProteins and enzymes can be digested by
adding a protease to the solution Proteases break down proteins into small peptides and amino acids
46
PRECIPITATING AND PURIFYING Precipitate DNA
Adding cold alcohol to a solution will cause DNA to precipitate and aggregate The DNA can be collected by centrifuging the sample and pouring off the liquid layer The DNA should exist as a small pellet in the bottom of the centrifuge tube
Purify DNAThe DNA can be washed by re-suspending it
in a cold alcohol solution and re-centrifuging it several times to obtain a very pure DNA sample Typical alcohols used to precipitate DNA include ethanol and Isopropanol This process leaves a very pure sample for stringent DNA testing
47
PART B OBSERVING EXTRACTED DNA UNDER THE MICROSCOPE
48
PART B A CLOSER LOOK
8 Use a dropping pipette to carefully remove some of the threadlike substance from the top of your preparation
9 Place one or two drops onto the middle of a
microscope slide
10 Add two drops of methylene blue Wait 3 or 4 minutes to allow the methylene blue to be absorbed by the DNA
11 Carefully place a cover slip on the slide Gently press
a folded piece of paper towelling over the top of the prepared slide to soak up any excess liquid
12 Observe the DNA under low power then high power
49
DISCUSSIONANSWER IN GROUP RELATED WORKED 1 Write a detailed description of the material
floating at the top of the test tube after the alcohol was added
2 Describe the DNA as it appears under the
microscope under high power 3 Prepare a diagram of your DNA specimen 4 What was the reason for using methylene
blue
50
HOMEWORKCONSTRUCT A FLOW CHART DRAWING MIND MAP OF THE PROCESS YOU USED EXTRACTING THE DNA FROM KIWIrsquoS
Next to each step explain how the method you used was important (Hint What substances make up the membranes of cells and cell organelles
How do detergents work
What is the active ingredient in meat tenderiser
51
TIPS STEPS FOR FLOW CHART In order to release the DNA from the
nuclei of the pea cells you first separated the cells from one another
Then the cell and nuclear membranes needed to be ruptured to release the cell contents and the contents of the nucleus
Once removed from the nuclear membrane the DNA had to be untangled into the visible threadlike structures you ended up with
52
HOMEWORK EXTENSION GROUP QUESTIONS Where can DNA be found in the cell Discuss the action of the soap (detergent) on the cell
What is the purpose of the soap in this activity What was the purpose of the Sodium Chloride Include a
discussion of polarity and charged particles Why was the cold ethanol added to the soap and salt
mixture Describe the appearance of your final product Draw a diagram of DNA containing 5 sets of nucleotide
bases labelling the hydrogen bonds between the bases References and Resources Adapted from Berry Full
of DNA by Diane Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
53
HTTPWWWGENOMEBCCAEDUCATIONTEACHERSCLASSROOM-ACTIVITIESDNA-EXTRACTION
54
EXTRACHROMOSOMES EXIST IN PAIRS Because our chromosomes exist in pairs (and consequently we
have 2 alleles of each gene) we are a diploid species This is why our somatic cells are represented as 2n Our gametes (sperm and ova) on the other hand are haploid and are represented as n Other species may have different ploidy for example
triploid (3n) seedless watermelons tetaploid (4n) salmonidae fish pentaploid (5n) Kenai birch hexaploid (6n) some types of wheat kiwi fruit octaploid (8n) acipenser (a genus
of sturgeon fish strawberies) decaploid (10n) some strawberries dodecaploid (12n) some types of amphibians eg Xenopus
ruwenzoriensis
wwwcyberusca
55
YOUR EXTRACTED DNA UNDER THE MICROSCOPE LOOKED
LIKE
Collaboration-Brainstorming on what to look at under the
- DNA extraction- DNA is too small for even a microscope to see and after the extraction it just appears like a blob True or false
Could you any DNA strand
wwwemployeescsbsjuedu
56
HANDOUT ON DNA EXTRACTIONBACKGROUND EVERY DAY LIFE
EXTRA READING MATERIAL(EXTENSION )
DISCOVERING DNA DNA ON THE INSIDE USE OF DNA EXTRACTION INN EVERYDAY LIFE WHY IS DNA TESTING GOOD MOLECULAR BIOLOGY PCR-based diagnostics of genomic DNA
57
REFERENCES httpwwwsquidoocomhow-to-get-dna-from-a-kiwi-fruit Why Is DNA Testing Good | eHowcom
httpwwwehowcomabout_6684410_dna-testing-good_htmlixzz1MkbcIJs5 Why Is DNA Extraction Important | eHowcom
httpwwwehowcomlist_5839095_dna-extraction-important_htmlixzz1MkZDrqyY
Why Is Sodium Used in DNA Extraction | eHowcom httpwwwehowcomabout_6504902_sodium-used-dna-extraction_htmlixzz1MkaGWg57
Why Is Cold Alcohol Used in DNA Tests | eHowcom httpwwwehowcomabout_6399349_cold-alcohol-used-dna-tests_htmlixzz1MkbEmEdu
httplearngeneticsutaheducontentlabsextractionhowto www kamibudaksainsblogspotcom
httpwwwchemwatch goldcom References and Resources Adapted from Berry Full of DNA by Diane
Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
58
REFERENCES
Uses of DNA Extraction | eHowcom httpwwwehowcomabout_5344428_uses-dna-extractionhtmlixzz1MkWQB2TZ
Science 10 A Contextual Approach Heineman Queensland Science Projects Regan Spence Maggie Spenceley
httpenwikipediaorgwikiMolecular_biology httpwwwclinchemorgcgicontent
extract44102201 http
employeescsbsjueduhjakubowskiclassesch331dnaoldnastructurehtml
httpwwwgenomebccaeducationteachersclassroom-activitiesdna-extraction
21
WHAT MUST I DO NOWMEAT TENDERIZER
ENZYME POWDERALCOHOL SEPARATION
wwwforumsoverclockerscomau
22
4 Add a pinch of enzyme (meat tenderizer) to mixture
(Using pineapple juice or contact lens cleaning solution will do the same as tenderizer)
Gently stir with toothpick skewer for 5 minutes continue stirring not too vigorously Be careful If you stir too hard youll break up the DNA making it harder to see
5 Quarter-fill a large test tube with the mixture
23
SLOWLY pour the same amount ice - cold (70 - 95 isopropyl or ethyl alcohol) into test tube pour alcohol down the side of the test tube
Tilting the test tube will make this easier to do
It forms a layer on top of the cell mixture
DO NOT MIX THE TWO LAYERS TOGETHER
Amount of alcohol and mixture should be the same
24
SEPARATION USING ALCOHOL
25
7 Observe the mixture for a few minutes You will see a white threadlike substance rise from the mixture to rest above in the alcohol layer
This is the DNA that you have extracted from the cells of the kiwi fruit
8 You can get more DNA to precipitate from the solution using a DNA collecting tool (glass or paper clip hook or cut inoculation needle)
Gently lift the water solution up into the alcohol layer (this allows more DNA to get in contact with the alcohol and precipitate)
26
INFORMATION DNA precipitates as a white stringy
ldquosnottyrdquo film at the water - alcohol interface and eventually will rise into the alcohol layer from the mixture layer
Allow test tube to sit for several minutes The clearer the DNA is the fewer
impurities you have
If you have an acceptable amount of DNA it can be spooled by rotating your collecting tool and then transferred into a clean tube container
27
WOW ndash SPOOLING THE DNA If you are careful you may be able wind up the DNA
around a glass rod or a skewer Position the tip of the glass rod or skewer where you can see the threads of DNA Steadily twist the rod or skewer as if you were making candy floss Alternatively use a straw to pull it out by suction ndash be careful not to get in you mouth
Donrsquot go too quickly
You should be able to pull the strands of DNA out of the mixture
ASK STUDENTS TO TAKE MOBILE PHONE PICTURES OF THEIR OWN EXTRACTED DNA AND COMPARE APPEARANCE WITH OTHERS
Photos of steps can be inserted in flow diagram(ICT)
28
KIWIDNA
29
If you want to save your DNA you can transfer it to a small container filled with alcohol
Leave tube container uncapped until the ethanol has evaporated
DNA can be stored in the fridge (dry or water buffer can be added)
You can now investigate the property
30
WHAT IS THAT STRINGY STUFFDNA IS A LONG STRINGY MOLECULE THE SALT THAT YOU ADDED IN STEP ONE HELPS IT STICK TOGETHER SO WHAT YOU SEE ARE CLUMPS OF TANGLED DNA MOLECULES
DNA NORMALLY STAYS DISSOLVED IN WATER BUT WHEN SALTY DNA COMES IN CONTACT WITH ALCOHOL IT BECOMES UNDISSOLVED THIS IS CALLED PRECIPITATION THE PHYSICAL FORCE OF THE DNA CLUMPING TOGETHER AS IT PRECIPITATES PULLS MORE STRANDS ALONG WITH IT AS IT RISES INTO THE ALCOHOL
31
THE WHYrsquoS Blending separated the pea cells In order to extract DNA from a cell the associated
membranes and proteins must first be removed (break apart the cells) and then physically separated (loosen the tough cell wall) from the DNA
To see the DNA we have to break open these two sacks We do this with detergent and salt( Sodium can be
involved in several of the steps ) Sodium is an element Its chemical symbol is Na for
Natrium the Latin word for sodium It is a positive ion and often associates with negative ions as part of useful compounds
Salt help precipitate protein and carbohydrates away from the DNA
Salt helps strip away the proteins associated with DNA + and ndash charge
32
THE WHYrsquoS Salt and Detergents are used to break down
cell walls and nuclear membranes to release the DNA
They work by chemically poking holes in the cell membranes or walls
Once holes are poked in the membranes the membranes can be further disrupted mechanically as with a blender
After that it is easier to get the contents of the cell out including the DNA
LETS HAVE A LOOK AT THE CELL
33
CELL TO DNA
httplearngeneticsutaheducontentlabsextractionhowtoHOW TO EXTRACT DNA FROM ANYTHING LIVING
34
THE WHYrsquoS WHY DETERGENT
Each cell is surrounded by a sack (the cell membrane)
DNA is found inside the second sack (nucleus) within each cell
To see the DNA we have to break it open A cell membrane has 2 layers of lipid(fat)
molecules with protein going through them When the lysis buffer (detergent) comes
close to the cell it captures the lipids and the proteins - breaks open the cell destroying the fatty membrane - DNA now released into the solution
35
A CELLS MEMBRANES HAVE TWO LAYERS OF LIPID (FAT) MOLECULES WITH PROTEINS GOING THROUGH THEM
36
Why detergent How does detergent work
Think about why you use soap to wash dishes or your hands To remove grease and dirt right
Soap molecules and grease molecules are made of two parts
1(Blue) Heads which like water 2(Green) Tails which hate water
37
AFTER ADDING THE DETERGENT WHAT DO YOU HAVE IN YOUR SOUP
38
THE WHYrsquoS ENZYME (MEAT TENDERIZER)
The DNA in the nucleus of the cell is molded folded and protected by proteins The tenderizer cut the proteins away from the DNA
In this experiment meat tenderizer acts as an enzyme to cut proteins just like a pair of scissors
The meat tenderizer cuts the proteins away from the DNA
39
THE WHYrsquoS CUTTING PROTEIN AND DNA
The DNA in the nucleus of the cell is moulded folded and protected by proteins
40
THE WHYrsquoSALCOHOL Alcohol is less dense than water so it floats
on top Look for clumps of white stringy stuff where the water and alcohol layers meet
DNA is not soluble in alcohol - other cell parts are
By adding alcohol DNA precipitates out of the solution and collect at the interface of the alcohol and soap layer
The colder the alcohol the less soluble the DNA will be in it
41
COLD ALCOHOL
DNA dissolves in water but precipitates in alcohol
Cold alcohol is used to separate DNA out of water-based solutions
This allows the DNA to be purified for subsequent genetic testing
Adding alcohol to a solution containing DNA is a simple way to obtain the pure DNA required and colder temperatures slow down enzymes that can break down DNA giving better extraction results
42
WHY IS DNA EXTRACTION IMPORTANT DNA extraction is an important molecular
biology procedure
By definition extraction is taking DNA out of any type of cell for the purpose of analysis
See Handouts for more information on the use of DNA extracted (extension only)
43
SUMMARIZEDNA EXTRACTING FROM KIWI FRUIT
44
BREAK DOWN CELL WALLS
Cells walls have to be destroyed to reach the DNA within cells
Extraction procedures to obtain pure DNA have to get rid of all the molecular and chemical components of the tissue from which the DNA is being extracted
First steps involve lysing or destroying the cell walls This can be done with a variety of chemical agents that are caustic to cell membranes but do not harm the DNA Cells can also be sonicated homogenized or ground up to destroy membranes
45
REMOVING Removing Lipids
Once the cell walls have been destroyed a detergent is added to get rid of the fats and oils that make up the cell membranes Detergents cause the fats and oils to dissolve into the solution
Remove ProteinsProteins and enzymes can be digested by
adding a protease to the solution Proteases break down proteins into small peptides and amino acids
46
PRECIPITATING AND PURIFYING Precipitate DNA
Adding cold alcohol to a solution will cause DNA to precipitate and aggregate The DNA can be collected by centrifuging the sample and pouring off the liquid layer The DNA should exist as a small pellet in the bottom of the centrifuge tube
Purify DNAThe DNA can be washed by re-suspending it
in a cold alcohol solution and re-centrifuging it several times to obtain a very pure DNA sample Typical alcohols used to precipitate DNA include ethanol and Isopropanol This process leaves a very pure sample for stringent DNA testing
47
PART B OBSERVING EXTRACTED DNA UNDER THE MICROSCOPE
48
PART B A CLOSER LOOK
8 Use a dropping pipette to carefully remove some of the threadlike substance from the top of your preparation
9 Place one or two drops onto the middle of a
microscope slide
10 Add two drops of methylene blue Wait 3 or 4 minutes to allow the methylene blue to be absorbed by the DNA
11 Carefully place a cover slip on the slide Gently press
a folded piece of paper towelling over the top of the prepared slide to soak up any excess liquid
12 Observe the DNA under low power then high power
49
DISCUSSIONANSWER IN GROUP RELATED WORKED 1 Write a detailed description of the material
floating at the top of the test tube after the alcohol was added
2 Describe the DNA as it appears under the
microscope under high power 3 Prepare a diagram of your DNA specimen 4 What was the reason for using methylene
blue
50
HOMEWORKCONSTRUCT A FLOW CHART DRAWING MIND MAP OF THE PROCESS YOU USED EXTRACTING THE DNA FROM KIWIrsquoS
Next to each step explain how the method you used was important (Hint What substances make up the membranes of cells and cell organelles
How do detergents work
What is the active ingredient in meat tenderiser
51
TIPS STEPS FOR FLOW CHART In order to release the DNA from the
nuclei of the pea cells you first separated the cells from one another
Then the cell and nuclear membranes needed to be ruptured to release the cell contents and the contents of the nucleus
Once removed from the nuclear membrane the DNA had to be untangled into the visible threadlike structures you ended up with
52
HOMEWORK EXTENSION GROUP QUESTIONS Where can DNA be found in the cell Discuss the action of the soap (detergent) on the cell
What is the purpose of the soap in this activity What was the purpose of the Sodium Chloride Include a
discussion of polarity and charged particles Why was the cold ethanol added to the soap and salt
mixture Describe the appearance of your final product Draw a diagram of DNA containing 5 sets of nucleotide
bases labelling the hydrogen bonds between the bases References and Resources Adapted from Berry Full
of DNA by Diane Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
53
HTTPWWWGENOMEBCCAEDUCATIONTEACHERSCLASSROOM-ACTIVITIESDNA-EXTRACTION
54
EXTRACHROMOSOMES EXIST IN PAIRS Because our chromosomes exist in pairs (and consequently we
have 2 alleles of each gene) we are a diploid species This is why our somatic cells are represented as 2n Our gametes (sperm and ova) on the other hand are haploid and are represented as n Other species may have different ploidy for example
triploid (3n) seedless watermelons tetaploid (4n) salmonidae fish pentaploid (5n) Kenai birch hexaploid (6n) some types of wheat kiwi fruit octaploid (8n) acipenser (a genus
of sturgeon fish strawberies) decaploid (10n) some strawberries dodecaploid (12n) some types of amphibians eg Xenopus
ruwenzoriensis
wwwcyberusca
55
YOUR EXTRACTED DNA UNDER THE MICROSCOPE LOOKED
LIKE
Collaboration-Brainstorming on what to look at under the
- DNA extraction- DNA is too small for even a microscope to see and after the extraction it just appears like a blob True or false
Could you any DNA strand
wwwemployeescsbsjuedu
56
HANDOUT ON DNA EXTRACTIONBACKGROUND EVERY DAY LIFE
EXTRA READING MATERIAL(EXTENSION )
DISCOVERING DNA DNA ON THE INSIDE USE OF DNA EXTRACTION INN EVERYDAY LIFE WHY IS DNA TESTING GOOD MOLECULAR BIOLOGY PCR-based diagnostics of genomic DNA
57
REFERENCES httpwwwsquidoocomhow-to-get-dna-from-a-kiwi-fruit Why Is DNA Testing Good | eHowcom
httpwwwehowcomabout_6684410_dna-testing-good_htmlixzz1MkbcIJs5 Why Is DNA Extraction Important | eHowcom
httpwwwehowcomlist_5839095_dna-extraction-important_htmlixzz1MkZDrqyY
Why Is Sodium Used in DNA Extraction | eHowcom httpwwwehowcomabout_6504902_sodium-used-dna-extraction_htmlixzz1MkaGWg57
Why Is Cold Alcohol Used in DNA Tests | eHowcom httpwwwehowcomabout_6399349_cold-alcohol-used-dna-tests_htmlixzz1MkbEmEdu
httplearngeneticsutaheducontentlabsextractionhowto www kamibudaksainsblogspotcom
httpwwwchemwatch goldcom References and Resources Adapted from Berry Full of DNA by Diane
Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
58
REFERENCES
Uses of DNA Extraction | eHowcom httpwwwehowcomabout_5344428_uses-dna-extractionhtmlixzz1MkWQB2TZ
Science 10 A Contextual Approach Heineman Queensland Science Projects Regan Spence Maggie Spenceley
httpenwikipediaorgwikiMolecular_biology httpwwwclinchemorgcgicontent
extract44102201 http
employeescsbsjueduhjakubowskiclassesch331dnaoldnastructurehtml
httpwwwgenomebccaeducationteachersclassroom-activitiesdna-extraction
22
4 Add a pinch of enzyme (meat tenderizer) to mixture
(Using pineapple juice or contact lens cleaning solution will do the same as tenderizer)
Gently stir with toothpick skewer for 5 minutes continue stirring not too vigorously Be careful If you stir too hard youll break up the DNA making it harder to see
5 Quarter-fill a large test tube with the mixture
23
SLOWLY pour the same amount ice - cold (70 - 95 isopropyl or ethyl alcohol) into test tube pour alcohol down the side of the test tube
Tilting the test tube will make this easier to do
It forms a layer on top of the cell mixture
DO NOT MIX THE TWO LAYERS TOGETHER
Amount of alcohol and mixture should be the same
24
SEPARATION USING ALCOHOL
25
7 Observe the mixture for a few minutes You will see a white threadlike substance rise from the mixture to rest above in the alcohol layer
This is the DNA that you have extracted from the cells of the kiwi fruit
8 You can get more DNA to precipitate from the solution using a DNA collecting tool (glass or paper clip hook or cut inoculation needle)
Gently lift the water solution up into the alcohol layer (this allows more DNA to get in contact with the alcohol and precipitate)
26
INFORMATION DNA precipitates as a white stringy
ldquosnottyrdquo film at the water - alcohol interface and eventually will rise into the alcohol layer from the mixture layer
Allow test tube to sit for several minutes The clearer the DNA is the fewer
impurities you have
If you have an acceptable amount of DNA it can be spooled by rotating your collecting tool and then transferred into a clean tube container
27
WOW ndash SPOOLING THE DNA If you are careful you may be able wind up the DNA
around a glass rod or a skewer Position the tip of the glass rod or skewer where you can see the threads of DNA Steadily twist the rod or skewer as if you were making candy floss Alternatively use a straw to pull it out by suction ndash be careful not to get in you mouth
Donrsquot go too quickly
You should be able to pull the strands of DNA out of the mixture
ASK STUDENTS TO TAKE MOBILE PHONE PICTURES OF THEIR OWN EXTRACTED DNA AND COMPARE APPEARANCE WITH OTHERS
Photos of steps can be inserted in flow diagram(ICT)
28
KIWIDNA
29
If you want to save your DNA you can transfer it to a small container filled with alcohol
Leave tube container uncapped until the ethanol has evaporated
DNA can be stored in the fridge (dry or water buffer can be added)
You can now investigate the property
30
WHAT IS THAT STRINGY STUFFDNA IS A LONG STRINGY MOLECULE THE SALT THAT YOU ADDED IN STEP ONE HELPS IT STICK TOGETHER SO WHAT YOU SEE ARE CLUMPS OF TANGLED DNA MOLECULES
DNA NORMALLY STAYS DISSOLVED IN WATER BUT WHEN SALTY DNA COMES IN CONTACT WITH ALCOHOL IT BECOMES UNDISSOLVED THIS IS CALLED PRECIPITATION THE PHYSICAL FORCE OF THE DNA CLUMPING TOGETHER AS IT PRECIPITATES PULLS MORE STRANDS ALONG WITH IT AS IT RISES INTO THE ALCOHOL
31
THE WHYrsquoS Blending separated the pea cells In order to extract DNA from a cell the associated
membranes and proteins must first be removed (break apart the cells) and then physically separated (loosen the tough cell wall) from the DNA
To see the DNA we have to break open these two sacks We do this with detergent and salt( Sodium can be
involved in several of the steps ) Sodium is an element Its chemical symbol is Na for
Natrium the Latin word for sodium It is a positive ion and often associates with negative ions as part of useful compounds
Salt help precipitate protein and carbohydrates away from the DNA
Salt helps strip away the proteins associated with DNA + and ndash charge
32
THE WHYrsquoS Salt and Detergents are used to break down
cell walls and nuclear membranes to release the DNA
They work by chemically poking holes in the cell membranes or walls
Once holes are poked in the membranes the membranes can be further disrupted mechanically as with a blender
After that it is easier to get the contents of the cell out including the DNA
LETS HAVE A LOOK AT THE CELL
33
CELL TO DNA
httplearngeneticsutaheducontentlabsextractionhowtoHOW TO EXTRACT DNA FROM ANYTHING LIVING
34
THE WHYrsquoS WHY DETERGENT
Each cell is surrounded by a sack (the cell membrane)
DNA is found inside the second sack (nucleus) within each cell
To see the DNA we have to break it open A cell membrane has 2 layers of lipid(fat)
molecules with protein going through them When the lysis buffer (detergent) comes
close to the cell it captures the lipids and the proteins - breaks open the cell destroying the fatty membrane - DNA now released into the solution
35
A CELLS MEMBRANES HAVE TWO LAYERS OF LIPID (FAT) MOLECULES WITH PROTEINS GOING THROUGH THEM
36
Why detergent How does detergent work
Think about why you use soap to wash dishes or your hands To remove grease and dirt right
Soap molecules and grease molecules are made of two parts
1(Blue) Heads which like water 2(Green) Tails which hate water
37
AFTER ADDING THE DETERGENT WHAT DO YOU HAVE IN YOUR SOUP
38
THE WHYrsquoS ENZYME (MEAT TENDERIZER)
The DNA in the nucleus of the cell is molded folded and protected by proteins The tenderizer cut the proteins away from the DNA
In this experiment meat tenderizer acts as an enzyme to cut proteins just like a pair of scissors
The meat tenderizer cuts the proteins away from the DNA
39
THE WHYrsquoS CUTTING PROTEIN AND DNA
The DNA in the nucleus of the cell is moulded folded and protected by proteins
40
THE WHYrsquoSALCOHOL Alcohol is less dense than water so it floats
on top Look for clumps of white stringy stuff where the water and alcohol layers meet
DNA is not soluble in alcohol - other cell parts are
By adding alcohol DNA precipitates out of the solution and collect at the interface of the alcohol and soap layer
The colder the alcohol the less soluble the DNA will be in it
41
COLD ALCOHOL
DNA dissolves in water but precipitates in alcohol
Cold alcohol is used to separate DNA out of water-based solutions
This allows the DNA to be purified for subsequent genetic testing
Adding alcohol to a solution containing DNA is a simple way to obtain the pure DNA required and colder temperatures slow down enzymes that can break down DNA giving better extraction results
42
WHY IS DNA EXTRACTION IMPORTANT DNA extraction is an important molecular
biology procedure
By definition extraction is taking DNA out of any type of cell for the purpose of analysis
See Handouts for more information on the use of DNA extracted (extension only)
43
SUMMARIZEDNA EXTRACTING FROM KIWI FRUIT
44
BREAK DOWN CELL WALLS
Cells walls have to be destroyed to reach the DNA within cells
Extraction procedures to obtain pure DNA have to get rid of all the molecular and chemical components of the tissue from which the DNA is being extracted
First steps involve lysing or destroying the cell walls This can be done with a variety of chemical agents that are caustic to cell membranes but do not harm the DNA Cells can also be sonicated homogenized or ground up to destroy membranes
45
REMOVING Removing Lipids
Once the cell walls have been destroyed a detergent is added to get rid of the fats and oils that make up the cell membranes Detergents cause the fats and oils to dissolve into the solution
Remove ProteinsProteins and enzymes can be digested by
adding a protease to the solution Proteases break down proteins into small peptides and amino acids
46
PRECIPITATING AND PURIFYING Precipitate DNA
Adding cold alcohol to a solution will cause DNA to precipitate and aggregate The DNA can be collected by centrifuging the sample and pouring off the liquid layer The DNA should exist as a small pellet in the bottom of the centrifuge tube
Purify DNAThe DNA can be washed by re-suspending it
in a cold alcohol solution and re-centrifuging it several times to obtain a very pure DNA sample Typical alcohols used to precipitate DNA include ethanol and Isopropanol This process leaves a very pure sample for stringent DNA testing
47
PART B OBSERVING EXTRACTED DNA UNDER THE MICROSCOPE
48
PART B A CLOSER LOOK
8 Use a dropping pipette to carefully remove some of the threadlike substance from the top of your preparation
9 Place one or two drops onto the middle of a
microscope slide
10 Add two drops of methylene blue Wait 3 or 4 minutes to allow the methylene blue to be absorbed by the DNA
11 Carefully place a cover slip on the slide Gently press
a folded piece of paper towelling over the top of the prepared slide to soak up any excess liquid
12 Observe the DNA under low power then high power
49
DISCUSSIONANSWER IN GROUP RELATED WORKED 1 Write a detailed description of the material
floating at the top of the test tube after the alcohol was added
2 Describe the DNA as it appears under the
microscope under high power 3 Prepare a diagram of your DNA specimen 4 What was the reason for using methylene
blue
50
HOMEWORKCONSTRUCT A FLOW CHART DRAWING MIND MAP OF THE PROCESS YOU USED EXTRACTING THE DNA FROM KIWIrsquoS
Next to each step explain how the method you used was important (Hint What substances make up the membranes of cells and cell organelles
How do detergents work
What is the active ingredient in meat tenderiser
51
TIPS STEPS FOR FLOW CHART In order to release the DNA from the
nuclei of the pea cells you first separated the cells from one another
Then the cell and nuclear membranes needed to be ruptured to release the cell contents and the contents of the nucleus
Once removed from the nuclear membrane the DNA had to be untangled into the visible threadlike structures you ended up with
52
HOMEWORK EXTENSION GROUP QUESTIONS Where can DNA be found in the cell Discuss the action of the soap (detergent) on the cell
What is the purpose of the soap in this activity What was the purpose of the Sodium Chloride Include a
discussion of polarity and charged particles Why was the cold ethanol added to the soap and salt
mixture Describe the appearance of your final product Draw a diagram of DNA containing 5 sets of nucleotide
bases labelling the hydrogen bonds between the bases References and Resources Adapted from Berry Full
of DNA by Diane Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
53
HTTPWWWGENOMEBCCAEDUCATIONTEACHERSCLASSROOM-ACTIVITIESDNA-EXTRACTION
54
EXTRACHROMOSOMES EXIST IN PAIRS Because our chromosomes exist in pairs (and consequently we
have 2 alleles of each gene) we are a diploid species This is why our somatic cells are represented as 2n Our gametes (sperm and ova) on the other hand are haploid and are represented as n Other species may have different ploidy for example
triploid (3n) seedless watermelons tetaploid (4n) salmonidae fish pentaploid (5n) Kenai birch hexaploid (6n) some types of wheat kiwi fruit octaploid (8n) acipenser (a genus
of sturgeon fish strawberies) decaploid (10n) some strawberries dodecaploid (12n) some types of amphibians eg Xenopus
ruwenzoriensis
wwwcyberusca
55
YOUR EXTRACTED DNA UNDER THE MICROSCOPE LOOKED
LIKE
Collaboration-Brainstorming on what to look at under the
- DNA extraction- DNA is too small for even a microscope to see and after the extraction it just appears like a blob True or false
Could you any DNA strand
wwwemployeescsbsjuedu
56
HANDOUT ON DNA EXTRACTIONBACKGROUND EVERY DAY LIFE
EXTRA READING MATERIAL(EXTENSION )
DISCOVERING DNA DNA ON THE INSIDE USE OF DNA EXTRACTION INN EVERYDAY LIFE WHY IS DNA TESTING GOOD MOLECULAR BIOLOGY PCR-based diagnostics of genomic DNA
57
REFERENCES httpwwwsquidoocomhow-to-get-dna-from-a-kiwi-fruit Why Is DNA Testing Good | eHowcom
httpwwwehowcomabout_6684410_dna-testing-good_htmlixzz1MkbcIJs5 Why Is DNA Extraction Important | eHowcom
httpwwwehowcomlist_5839095_dna-extraction-important_htmlixzz1MkZDrqyY
Why Is Sodium Used in DNA Extraction | eHowcom httpwwwehowcomabout_6504902_sodium-used-dna-extraction_htmlixzz1MkaGWg57
Why Is Cold Alcohol Used in DNA Tests | eHowcom httpwwwehowcomabout_6399349_cold-alcohol-used-dna-tests_htmlixzz1MkbEmEdu
httplearngeneticsutaheducontentlabsextractionhowto www kamibudaksainsblogspotcom
httpwwwchemwatch goldcom References and Resources Adapted from Berry Full of DNA by Diane
Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
58
REFERENCES
Uses of DNA Extraction | eHowcom httpwwwehowcomabout_5344428_uses-dna-extractionhtmlixzz1MkWQB2TZ
Science 10 A Contextual Approach Heineman Queensland Science Projects Regan Spence Maggie Spenceley
httpenwikipediaorgwikiMolecular_biology httpwwwclinchemorgcgicontent
extract44102201 http
employeescsbsjueduhjakubowskiclassesch331dnaoldnastructurehtml
httpwwwgenomebccaeducationteachersclassroom-activitiesdna-extraction
23
SLOWLY pour the same amount ice - cold (70 - 95 isopropyl or ethyl alcohol) into test tube pour alcohol down the side of the test tube
Tilting the test tube will make this easier to do
It forms a layer on top of the cell mixture
DO NOT MIX THE TWO LAYERS TOGETHER
Amount of alcohol and mixture should be the same
24
SEPARATION USING ALCOHOL
25
7 Observe the mixture for a few minutes You will see a white threadlike substance rise from the mixture to rest above in the alcohol layer
This is the DNA that you have extracted from the cells of the kiwi fruit
8 You can get more DNA to precipitate from the solution using a DNA collecting tool (glass or paper clip hook or cut inoculation needle)
Gently lift the water solution up into the alcohol layer (this allows more DNA to get in contact with the alcohol and precipitate)
26
INFORMATION DNA precipitates as a white stringy
ldquosnottyrdquo film at the water - alcohol interface and eventually will rise into the alcohol layer from the mixture layer
Allow test tube to sit for several minutes The clearer the DNA is the fewer
impurities you have
If you have an acceptable amount of DNA it can be spooled by rotating your collecting tool and then transferred into a clean tube container
27
WOW ndash SPOOLING THE DNA If you are careful you may be able wind up the DNA
around a glass rod or a skewer Position the tip of the glass rod or skewer where you can see the threads of DNA Steadily twist the rod or skewer as if you were making candy floss Alternatively use a straw to pull it out by suction ndash be careful not to get in you mouth
Donrsquot go too quickly
You should be able to pull the strands of DNA out of the mixture
ASK STUDENTS TO TAKE MOBILE PHONE PICTURES OF THEIR OWN EXTRACTED DNA AND COMPARE APPEARANCE WITH OTHERS
Photos of steps can be inserted in flow diagram(ICT)
28
KIWIDNA
29
If you want to save your DNA you can transfer it to a small container filled with alcohol
Leave tube container uncapped until the ethanol has evaporated
DNA can be stored in the fridge (dry or water buffer can be added)
You can now investigate the property
30
WHAT IS THAT STRINGY STUFFDNA IS A LONG STRINGY MOLECULE THE SALT THAT YOU ADDED IN STEP ONE HELPS IT STICK TOGETHER SO WHAT YOU SEE ARE CLUMPS OF TANGLED DNA MOLECULES
DNA NORMALLY STAYS DISSOLVED IN WATER BUT WHEN SALTY DNA COMES IN CONTACT WITH ALCOHOL IT BECOMES UNDISSOLVED THIS IS CALLED PRECIPITATION THE PHYSICAL FORCE OF THE DNA CLUMPING TOGETHER AS IT PRECIPITATES PULLS MORE STRANDS ALONG WITH IT AS IT RISES INTO THE ALCOHOL
31
THE WHYrsquoS Blending separated the pea cells In order to extract DNA from a cell the associated
membranes and proteins must first be removed (break apart the cells) and then physically separated (loosen the tough cell wall) from the DNA
To see the DNA we have to break open these two sacks We do this with detergent and salt( Sodium can be
involved in several of the steps ) Sodium is an element Its chemical symbol is Na for
Natrium the Latin word for sodium It is a positive ion and often associates with negative ions as part of useful compounds
Salt help precipitate protein and carbohydrates away from the DNA
Salt helps strip away the proteins associated with DNA + and ndash charge
32
THE WHYrsquoS Salt and Detergents are used to break down
cell walls and nuclear membranes to release the DNA
They work by chemically poking holes in the cell membranes or walls
Once holes are poked in the membranes the membranes can be further disrupted mechanically as with a blender
After that it is easier to get the contents of the cell out including the DNA
LETS HAVE A LOOK AT THE CELL
33
CELL TO DNA
httplearngeneticsutaheducontentlabsextractionhowtoHOW TO EXTRACT DNA FROM ANYTHING LIVING
34
THE WHYrsquoS WHY DETERGENT
Each cell is surrounded by a sack (the cell membrane)
DNA is found inside the second sack (nucleus) within each cell
To see the DNA we have to break it open A cell membrane has 2 layers of lipid(fat)
molecules with protein going through them When the lysis buffer (detergent) comes
close to the cell it captures the lipids and the proteins - breaks open the cell destroying the fatty membrane - DNA now released into the solution
35
A CELLS MEMBRANES HAVE TWO LAYERS OF LIPID (FAT) MOLECULES WITH PROTEINS GOING THROUGH THEM
36
Why detergent How does detergent work
Think about why you use soap to wash dishes or your hands To remove grease and dirt right
Soap molecules and grease molecules are made of two parts
1(Blue) Heads which like water 2(Green) Tails which hate water
37
AFTER ADDING THE DETERGENT WHAT DO YOU HAVE IN YOUR SOUP
38
THE WHYrsquoS ENZYME (MEAT TENDERIZER)
The DNA in the nucleus of the cell is molded folded and protected by proteins The tenderizer cut the proteins away from the DNA
In this experiment meat tenderizer acts as an enzyme to cut proteins just like a pair of scissors
The meat tenderizer cuts the proteins away from the DNA
39
THE WHYrsquoS CUTTING PROTEIN AND DNA
The DNA in the nucleus of the cell is moulded folded and protected by proteins
40
THE WHYrsquoSALCOHOL Alcohol is less dense than water so it floats
on top Look for clumps of white stringy stuff where the water and alcohol layers meet
DNA is not soluble in alcohol - other cell parts are
By adding alcohol DNA precipitates out of the solution and collect at the interface of the alcohol and soap layer
The colder the alcohol the less soluble the DNA will be in it
41
COLD ALCOHOL
DNA dissolves in water but precipitates in alcohol
Cold alcohol is used to separate DNA out of water-based solutions
This allows the DNA to be purified for subsequent genetic testing
Adding alcohol to a solution containing DNA is a simple way to obtain the pure DNA required and colder temperatures slow down enzymes that can break down DNA giving better extraction results
42
WHY IS DNA EXTRACTION IMPORTANT DNA extraction is an important molecular
biology procedure
By definition extraction is taking DNA out of any type of cell for the purpose of analysis
See Handouts for more information on the use of DNA extracted (extension only)
43
SUMMARIZEDNA EXTRACTING FROM KIWI FRUIT
44
BREAK DOWN CELL WALLS
Cells walls have to be destroyed to reach the DNA within cells
Extraction procedures to obtain pure DNA have to get rid of all the molecular and chemical components of the tissue from which the DNA is being extracted
First steps involve lysing or destroying the cell walls This can be done with a variety of chemical agents that are caustic to cell membranes but do not harm the DNA Cells can also be sonicated homogenized or ground up to destroy membranes
45
REMOVING Removing Lipids
Once the cell walls have been destroyed a detergent is added to get rid of the fats and oils that make up the cell membranes Detergents cause the fats and oils to dissolve into the solution
Remove ProteinsProteins and enzymes can be digested by
adding a protease to the solution Proteases break down proteins into small peptides and amino acids
46
PRECIPITATING AND PURIFYING Precipitate DNA
Adding cold alcohol to a solution will cause DNA to precipitate and aggregate The DNA can be collected by centrifuging the sample and pouring off the liquid layer The DNA should exist as a small pellet in the bottom of the centrifuge tube
Purify DNAThe DNA can be washed by re-suspending it
in a cold alcohol solution and re-centrifuging it several times to obtain a very pure DNA sample Typical alcohols used to precipitate DNA include ethanol and Isopropanol This process leaves a very pure sample for stringent DNA testing
47
PART B OBSERVING EXTRACTED DNA UNDER THE MICROSCOPE
48
PART B A CLOSER LOOK
8 Use a dropping pipette to carefully remove some of the threadlike substance from the top of your preparation
9 Place one or two drops onto the middle of a
microscope slide
10 Add two drops of methylene blue Wait 3 or 4 minutes to allow the methylene blue to be absorbed by the DNA
11 Carefully place a cover slip on the slide Gently press
a folded piece of paper towelling over the top of the prepared slide to soak up any excess liquid
12 Observe the DNA under low power then high power
49
DISCUSSIONANSWER IN GROUP RELATED WORKED 1 Write a detailed description of the material
floating at the top of the test tube after the alcohol was added
2 Describe the DNA as it appears under the
microscope under high power 3 Prepare a diagram of your DNA specimen 4 What was the reason for using methylene
blue
50
HOMEWORKCONSTRUCT A FLOW CHART DRAWING MIND MAP OF THE PROCESS YOU USED EXTRACTING THE DNA FROM KIWIrsquoS
Next to each step explain how the method you used was important (Hint What substances make up the membranes of cells and cell organelles
How do detergents work
What is the active ingredient in meat tenderiser
51
TIPS STEPS FOR FLOW CHART In order to release the DNA from the
nuclei of the pea cells you first separated the cells from one another
Then the cell and nuclear membranes needed to be ruptured to release the cell contents and the contents of the nucleus
Once removed from the nuclear membrane the DNA had to be untangled into the visible threadlike structures you ended up with
52
HOMEWORK EXTENSION GROUP QUESTIONS Where can DNA be found in the cell Discuss the action of the soap (detergent) on the cell
What is the purpose of the soap in this activity What was the purpose of the Sodium Chloride Include a
discussion of polarity and charged particles Why was the cold ethanol added to the soap and salt
mixture Describe the appearance of your final product Draw a diagram of DNA containing 5 sets of nucleotide
bases labelling the hydrogen bonds between the bases References and Resources Adapted from Berry Full
of DNA by Diane Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
53
HTTPWWWGENOMEBCCAEDUCATIONTEACHERSCLASSROOM-ACTIVITIESDNA-EXTRACTION
54
EXTRACHROMOSOMES EXIST IN PAIRS Because our chromosomes exist in pairs (and consequently we
have 2 alleles of each gene) we are a diploid species This is why our somatic cells are represented as 2n Our gametes (sperm and ova) on the other hand are haploid and are represented as n Other species may have different ploidy for example
triploid (3n) seedless watermelons tetaploid (4n) salmonidae fish pentaploid (5n) Kenai birch hexaploid (6n) some types of wheat kiwi fruit octaploid (8n) acipenser (a genus
of sturgeon fish strawberies) decaploid (10n) some strawberries dodecaploid (12n) some types of amphibians eg Xenopus
ruwenzoriensis
wwwcyberusca
55
YOUR EXTRACTED DNA UNDER THE MICROSCOPE LOOKED
LIKE
Collaboration-Brainstorming on what to look at under the
- DNA extraction- DNA is too small for even a microscope to see and after the extraction it just appears like a blob True or false
Could you any DNA strand
wwwemployeescsbsjuedu
56
HANDOUT ON DNA EXTRACTIONBACKGROUND EVERY DAY LIFE
EXTRA READING MATERIAL(EXTENSION )
DISCOVERING DNA DNA ON THE INSIDE USE OF DNA EXTRACTION INN EVERYDAY LIFE WHY IS DNA TESTING GOOD MOLECULAR BIOLOGY PCR-based diagnostics of genomic DNA
57
REFERENCES httpwwwsquidoocomhow-to-get-dna-from-a-kiwi-fruit Why Is DNA Testing Good | eHowcom
httpwwwehowcomabout_6684410_dna-testing-good_htmlixzz1MkbcIJs5 Why Is DNA Extraction Important | eHowcom
httpwwwehowcomlist_5839095_dna-extraction-important_htmlixzz1MkZDrqyY
Why Is Sodium Used in DNA Extraction | eHowcom httpwwwehowcomabout_6504902_sodium-used-dna-extraction_htmlixzz1MkaGWg57
Why Is Cold Alcohol Used in DNA Tests | eHowcom httpwwwehowcomabout_6399349_cold-alcohol-used-dna-tests_htmlixzz1MkbEmEdu
httplearngeneticsutaheducontentlabsextractionhowto www kamibudaksainsblogspotcom
httpwwwchemwatch goldcom References and Resources Adapted from Berry Full of DNA by Diane
Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
58
REFERENCES
Uses of DNA Extraction | eHowcom httpwwwehowcomabout_5344428_uses-dna-extractionhtmlixzz1MkWQB2TZ
Science 10 A Contextual Approach Heineman Queensland Science Projects Regan Spence Maggie Spenceley
httpenwikipediaorgwikiMolecular_biology httpwwwclinchemorgcgicontent
extract44102201 http
employeescsbsjueduhjakubowskiclassesch331dnaoldnastructurehtml
httpwwwgenomebccaeducationteachersclassroom-activitiesdna-extraction
24
SEPARATION USING ALCOHOL
25
7 Observe the mixture for a few minutes You will see a white threadlike substance rise from the mixture to rest above in the alcohol layer
This is the DNA that you have extracted from the cells of the kiwi fruit
8 You can get more DNA to precipitate from the solution using a DNA collecting tool (glass or paper clip hook or cut inoculation needle)
Gently lift the water solution up into the alcohol layer (this allows more DNA to get in contact with the alcohol and precipitate)
26
INFORMATION DNA precipitates as a white stringy
ldquosnottyrdquo film at the water - alcohol interface and eventually will rise into the alcohol layer from the mixture layer
Allow test tube to sit for several minutes The clearer the DNA is the fewer
impurities you have
If you have an acceptable amount of DNA it can be spooled by rotating your collecting tool and then transferred into a clean tube container
27
WOW ndash SPOOLING THE DNA If you are careful you may be able wind up the DNA
around a glass rod or a skewer Position the tip of the glass rod or skewer where you can see the threads of DNA Steadily twist the rod or skewer as if you were making candy floss Alternatively use a straw to pull it out by suction ndash be careful not to get in you mouth
Donrsquot go too quickly
You should be able to pull the strands of DNA out of the mixture
ASK STUDENTS TO TAKE MOBILE PHONE PICTURES OF THEIR OWN EXTRACTED DNA AND COMPARE APPEARANCE WITH OTHERS
Photos of steps can be inserted in flow diagram(ICT)
28
KIWIDNA
29
If you want to save your DNA you can transfer it to a small container filled with alcohol
Leave tube container uncapped until the ethanol has evaporated
DNA can be stored in the fridge (dry or water buffer can be added)
You can now investigate the property
30
WHAT IS THAT STRINGY STUFFDNA IS A LONG STRINGY MOLECULE THE SALT THAT YOU ADDED IN STEP ONE HELPS IT STICK TOGETHER SO WHAT YOU SEE ARE CLUMPS OF TANGLED DNA MOLECULES
DNA NORMALLY STAYS DISSOLVED IN WATER BUT WHEN SALTY DNA COMES IN CONTACT WITH ALCOHOL IT BECOMES UNDISSOLVED THIS IS CALLED PRECIPITATION THE PHYSICAL FORCE OF THE DNA CLUMPING TOGETHER AS IT PRECIPITATES PULLS MORE STRANDS ALONG WITH IT AS IT RISES INTO THE ALCOHOL
31
THE WHYrsquoS Blending separated the pea cells In order to extract DNA from a cell the associated
membranes and proteins must first be removed (break apart the cells) and then physically separated (loosen the tough cell wall) from the DNA
To see the DNA we have to break open these two sacks We do this with detergent and salt( Sodium can be
involved in several of the steps ) Sodium is an element Its chemical symbol is Na for
Natrium the Latin word for sodium It is a positive ion and often associates with negative ions as part of useful compounds
Salt help precipitate protein and carbohydrates away from the DNA
Salt helps strip away the proteins associated with DNA + and ndash charge
32
THE WHYrsquoS Salt and Detergents are used to break down
cell walls and nuclear membranes to release the DNA
They work by chemically poking holes in the cell membranes or walls
Once holes are poked in the membranes the membranes can be further disrupted mechanically as with a blender
After that it is easier to get the contents of the cell out including the DNA
LETS HAVE A LOOK AT THE CELL
33
CELL TO DNA
httplearngeneticsutaheducontentlabsextractionhowtoHOW TO EXTRACT DNA FROM ANYTHING LIVING
34
THE WHYrsquoS WHY DETERGENT
Each cell is surrounded by a sack (the cell membrane)
DNA is found inside the second sack (nucleus) within each cell
To see the DNA we have to break it open A cell membrane has 2 layers of lipid(fat)
molecules with protein going through them When the lysis buffer (detergent) comes
close to the cell it captures the lipids and the proteins - breaks open the cell destroying the fatty membrane - DNA now released into the solution
35
A CELLS MEMBRANES HAVE TWO LAYERS OF LIPID (FAT) MOLECULES WITH PROTEINS GOING THROUGH THEM
36
Why detergent How does detergent work
Think about why you use soap to wash dishes or your hands To remove grease and dirt right
Soap molecules and grease molecules are made of two parts
1(Blue) Heads which like water 2(Green) Tails which hate water
37
AFTER ADDING THE DETERGENT WHAT DO YOU HAVE IN YOUR SOUP
38
THE WHYrsquoS ENZYME (MEAT TENDERIZER)
The DNA in the nucleus of the cell is molded folded and protected by proteins The tenderizer cut the proteins away from the DNA
In this experiment meat tenderizer acts as an enzyme to cut proteins just like a pair of scissors
The meat tenderizer cuts the proteins away from the DNA
39
THE WHYrsquoS CUTTING PROTEIN AND DNA
The DNA in the nucleus of the cell is moulded folded and protected by proteins
40
THE WHYrsquoSALCOHOL Alcohol is less dense than water so it floats
on top Look for clumps of white stringy stuff where the water and alcohol layers meet
DNA is not soluble in alcohol - other cell parts are
By adding alcohol DNA precipitates out of the solution and collect at the interface of the alcohol and soap layer
The colder the alcohol the less soluble the DNA will be in it
41
COLD ALCOHOL
DNA dissolves in water but precipitates in alcohol
Cold alcohol is used to separate DNA out of water-based solutions
This allows the DNA to be purified for subsequent genetic testing
Adding alcohol to a solution containing DNA is a simple way to obtain the pure DNA required and colder temperatures slow down enzymes that can break down DNA giving better extraction results
42
WHY IS DNA EXTRACTION IMPORTANT DNA extraction is an important molecular
biology procedure
By definition extraction is taking DNA out of any type of cell for the purpose of analysis
See Handouts for more information on the use of DNA extracted (extension only)
43
SUMMARIZEDNA EXTRACTING FROM KIWI FRUIT
44
BREAK DOWN CELL WALLS
Cells walls have to be destroyed to reach the DNA within cells
Extraction procedures to obtain pure DNA have to get rid of all the molecular and chemical components of the tissue from which the DNA is being extracted
First steps involve lysing or destroying the cell walls This can be done with a variety of chemical agents that are caustic to cell membranes but do not harm the DNA Cells can also be sonicated homogenized or ground up to destroy membranes
45
REMOVING Removing Lipids
Once the cell walls have been destroyed a detergent is added to get rid of the fats and oils that make up the cell membranes Detergents cause the fats and oils to dissolve into the solution
Remove ProteinsProteins and enzymes can be digested by
adding a protease to the solution Proteases break down proteins into small peptides and amino acids
46
PRECIPITATING AND PURIFYING Precipitate DNA
Adding cold alcohol to a solution will cause DNA to precipitate and aggregate The DNA can be collected by centrifuging the sample and pouring off the liquid layer The DNA should exist as a small pellet in the bottom of the centrifuge tube
Purify DNAThe DNA can be washed by re-suspending it
in a cold alcohol solution and re-centrifuging it several times to obtain a very pure DNA sample Typical alcohols used to precipitate DNA include ethanol and Isopropanol This process leaves a very pure sample for stringent DNA testing
47
PART B OBSERVING EXTRACTED DNA UNDER THE MICROSCOPE
48
PART B A CLOSER LOOK
8 Use a dropping pipette to carefully remove some of the threadlike substance from the top of your preparation
9 Place one or two drops onto the middle of a
microscope slide
10 Add two drops of methylene blue Wait 3 or 4 minutes to allow the methylene blue to be absorbed by the DNA
11 Carefully place a cover slip on the slide Gently press
a folded piece of paper towelling over the top of the prepared slide to soak up any excess liquid
12 Observe the DNA under low power then high power
49
DISCUSSIONANSWER IN GROUP RELATED WORKED 1 Write a detailed description of the material
floating at the top of the test tube after the alcohol was added
2 Describe the DNA as it appears under the
microscope under high power 3 Prepare a diagram of your DNA specimen 4 What was the reason for using methylene
blue
50
HOMEWORKCONSTRUCT A FLOW CHART DRAWING MIND MAP OF THE PROCESS YOU USED EXTRACTING THE DNA FROM KIWIrsquoS
Next to each step explain how the method you used was important (Hint What substances make up the membranes of cells and cell organelles
How do detergents work
What is the active ingredient in meat tenderiser
51
TIPS STEPS FOR FLOW CHART In order to release the DNA from the
nuclei of the pea cells you first separated the cells from one another
Then the cell and nuclear membranes needed to be ruptured to release the cell contents and the contents of the nucleus
Once removed from the nuclear membrane the DNA had to be untangled into the visible threadlike structures you ended up with
52
HOMEWORK EXTENSION GROUP QUESTIONS Where can DNA be found in the cell Discuss the action of the soap (detergent) on the cell
What is the purpose of the soap in this activity What was the purpose of the Sodium Chloride Include a
discussion of polarity and charged particles Why was the cold ethanol added to the soap and salt
mixture Describe the appearance of your final product Draw a diagram of DNA containing 5 sets of nucleotide
bases labelling the hydrogen bonds between the bases References and Resources Adapted from Berry Full
of DNA by Diane Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
53
HTTPWWWGENOMEBCCAEDUCATIONTEACHERSCLASSROOM-ACTIVITIESDNA-EXTRACTION
54
EXTRACHROMOSOMES EXIST IN PAIRS Because our chromosomes exist in pairs (and consequently we
have 2 alleles of each gene) we are a diploid species This is why our somatic cells are represented as 2n Our gametes (sperm and ova) on the other hand are haploid and are represented as n Other species may have different ploidy for example
triploid (3n) seedless watermelons tetaploid (4n) salmonidae fish pentaploid (5n) Kenai birch hexaploid (6n) some types of wheat kiwi fruit octaploid (8n) acipenser (a genus
of sturgeon fish strawberies) decaploid (10n) some strawberries dodecaploid (12n) some types of amphibians eg Xenopus
ruwenzoriensis
wwwcyberusca
55
YOUR EXTRACTED DNA UNDER THE MICROSCOPE LOOKED
LIKE
Collaboration-Brainstorming on what to look at under the
- DNA extraction- DNA is too small for even a microscope to see and after the extraction it just appears like a blob True or false
Could you any DNA strand
wwwemployeescsbsjuedu
56
HANDOUT ON DNA EXTRACTIONBACKGROUND EVERY DAY LIFE
EXTRA READING MATERIAL(EXTENSION )
DISCOVERING DNA DNA ON THE INSIDE USE OF DNA EXTRACTION INN EVERYDAY LIFE WHY IS DNA TESTING GOOD MOLECULAR BIOLOGY PCR-based diagnostics of genomic DNA
57
REFERENCES httpwwwsquidoocomhow-to-get-dna-from-a-kiwi-fruit Why Is DNA Testing Good | eHowcom
httpwwwehowcomabout_6684410_dna-testing-good_htmlixzz1MkbcIJs5 Why Is DNA Extraction Important | eHowcom
httpwwwehowcomlist_5839095_dna-extraction-important_htmlixzz1MkZDrqyY
Why Is Sodium Used in DNA Extraction | eHowcom httpwwwehowcomabout_6504902_sodium-used-dna-extraction_htmlixzz1MkaGWg57
Why Is Cold Alcohol Used in DNA Tests | eHowcom httpwwwehowcomabout_6399349_cold-alcohol-used-dna-tests_htmlixzz1MkbEmEdu
httplearngeneticsutaheducontentlabsextractionhowto www kamibudaksainsblogspotcom
httpwwwchemwatch goldcom References and Resources Adapted from Berry Full of DNA by Diane
Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
58
REFERENCES
Uses of DNA Extraction | eHowcom httpwwwehowcomabout_5344428_uses-dna-extractionhtmlixzz1MkWQB2TZ
Science 10 A Contextual Approach Heineman Queensland Science Projects Regan Spence Maggie Spenceley
httpenwikipediaorgwikiMolecular_biology httpwwwclinchemorgcgicontent
extract44102201 http
employeescsbsjueduhjakubowskiclassesch331dnaoldnastructurehtml
httpwwwgenomebccaeducationteachersclassroom-activitiesdna-extraction
25
7 Observe the mixture for a few minutes You will see a white threadlike substance rise from the mixture to rest above in the alcohol layer
This is the DNA that you have extracted from the cells of the kiwi fruit
8 You can get more DNA to precipitate from the solution using a DNA collecting tool (glass or paper clip hook or cut inoculation needle)
Gently lift the water solution up into the alcohol layer (this allows more DNA to get in contact with the alcohol and precipitate)
26
INFORMATION DNA precipitates as a white stringy
ldquosnottyrdquo film at the water - alcohol interface and eventually will rise into the alcohol layer from the mixture layer
Allow test tube to sit for several minutes The clearer the DNA is the fewer
impurities you have
If you have an acceptable amount of DNA it can be spooled by rotating your collecting tool and then transferred into a clean tube container
27
WOW ndash SPOOLING THE DNA If you are careful you may be able wind up the DNA
around a glass rod or a skewer Position the tip of the glass rod or skewer where you can see the threads of DNA Steadily twist the rod or skewer as if you were making candy floss Alternatively use a straw to pull it out by suction ndash be careful not to get in you mouth
Donrsquot go too quickly
You should be able to pull the strands of DNA out of the mixture
ASK STUDENTS TO TAKE MOBILE PHONE PICTURES OF THEIR OWN EXTRACTED DNA AND COMPARE APPEARANCE WITH OTHERS
Photos of steps can be inserted in flow diagram(ICT)
28
KIWIDNA
29
If you want to save your DNA you can transfer it to a small container filled with alcohol
Leave tube container uncapped until the ethanol has evaporated
DNA can be stored in the fridge (dry or water buffer can be added)
You can now investigate the property
30
WHAT IS THAT STRINGY STUFFDNA IS A LONG STRINGY MOLECULE THE SALT THAT YOU ADDED IN STEP ONE HELPS IT STICK TOGETHER SO WHAT YOU SEE ARE CLUMPS OF TANGLED DNA MOLECULES
DNA NORMALLY STAYS DISSOLVED IN WATER BUT WHEN SALTY DNA COMES IN CONTACT WITH ALCOHOL IT BECOMES UNDISSOLVED THIS IS CALLED PRECIPITATION THE PHYSICAL FORCE OF THE DNA CLUMPING TOGETHER AS IT PRECIPITATES PULLS MORE STRANDS ALONG WITH IT AS IT RISES INTO THE ALCOHOL
31
THE WHYrsquoS Blending separated the pea cells In order to extract DNA from a cell the associated
membranes and proteins must first be removed (break apart the cells) and then physically separated (loosen the tough cell wall) from the DNA
To see the DNA we have to break open these two sacks We do this with detergent and salt( Sodium can be
involved in several of the steps ) Sodium is an element Its chemical symbol is Na for
Natrium the Latin word for sodium It is a positive ion and often associates with negative ions as part of useful compounds
Salt help precipitate protein and carbohydrates away from the DNA
Salt helps strip away the proteins associated with DNA + and ndash charge
32
THE WHYrsquoS Salt and Detergents are used to break down
cell walls and nuclear membranes to release the DNA
They work by chemically poking holes in the cell membranes or walls
Once holes are poked in the membranes the membranes can be further disrupted mechanically as with a blender
After that it is easier to get the contents of the cell out including the DNA
LETS HAVE A LOOK AT THE CELL
33
CELL TO DNA
httplearngeneticsutaheducontentlabsextractionhowtoHOW TO EXTRACT DNA FROM ANYTHING LIVING
34
THE WHYrsquoS WHY DETERGENT
Each cell is surrounded by a sack (the cell membrane)
DNA is found inside the second sack (nucleus) within each cell
To see the DNA we have to break it open A cell membrane has 2 layers of lipid(fat)
molecules with protein going through them When the lysis buffer (detergent) comes
close to the cell it captures the lipids and the proteins - breaks open the cell destroying the fatty membrane - DNA now released into the solution
35
A CELLS MEMBRANES HAVE TWO LAYERS OF LIPID (FAT) MOLECULES WITH PROTEINS GOING THROUGH THEM
36
Why detergent How does detergent work
Think about why you use soap to wash dishes or your hands To remove grease and dirt right
Soap molecules and grease molecules are made of two parts
1(Blue) Heads which like water 2(Green) Tails which hate water
37
AFTER ADDING THE DETERGENT WHAT DO YOU HAVE IN YOUR SOUP
38
THE WHYrsquoS ENZYME (MEAT TENDERIZER)
The DNA in the nucleus of the cell is molded folded and protected by proteins The tenderizer cut the proteins away from the DNA
In this experiment meat tenderizer acts as an enzyme to cut proteins just like a pair of scissors
The meat tenderizer cuts the proteins away from the DNA
39
THE WHYrsquoS CUTTING PROTEIN AND DNA
The DNA in the nucleus of the cell is moulded folded and protected by proteins
40
THE WHYrsquoSALCOHOL Alcohol is less dense than water so it floats
on top Look for clumps of white stringy stuff where the water and alcohol layers meet
DNA is not soluble in alcohol - other cell parts are
By adding alcohol DNA precipitates out of the solution and collect at the interface of the alcohol and soap layer
The colder the alcohol the less soluble the DNA will be in it
41
COLD ALCOHOL
DNA dissolves in water but precipitates in alcohol
Cold alcohol is used to separate DNA out of water-based solutions
This allows the DNA to be purified for subsequent genetic testing
Adding alcohol to a solution containing DNA is a simple way to obtain the pure DNA required and colder temperatures slow down enzymes that can break down DNA giving better extraction results
42
WHY IS DNA EXTRACTION IMPORTANT DNA extraction is an important molecular
biology procedure
By definition extraction is taking DNA out of any type of cell for the purpose of analysis
See Handouts for more information on the use of DNA extracted (extension only)
43
SUMMARIZEDNA EXTRACTING FROM KIWI FRUIT
44
BREAK DOWN CELL WALLS
Cells walls have to be destroyed to reach the DNA within cells
Extraction procedures to obtain pure DNA have to get rid of all the molecular and chemical components of the tissue from which the DNA is being extracted
First steps involve lysing or destroying the cell walls This can be done with a variety of chemical agents that are caustic to cell membranes but do not harm the DNA Cells can also be sonicated homogenized or ground up to destroy membranes
45
REMOVING Removing Lipids
Once the cell walls have been destroyed a detergent is added to get rid of the fats and oils that make up the cell membranes Detergents cause the fats and oils to dissolve into the solution
Remove ProteinsProteins and enzymes can be digested by
adding a protease to the solution Proteases break down proteins into small peptides and amino acids
46
PRECIPITATING AND PURIFYING Precipitate DNA
Adding cold alcohol to a solution will cause DNA to precipitate and aggregate The DNA can be collected by centrifuging the sample and pouring off the liquid layer The DNA should exist as a small pellet in the bottom of the centrifuge tube
Purify DNAThe DNA can be washed by re-suspending it
in a cold alcohol solution and re-centrifuging it several times to obtain a very pure DNA sample Typical alcohols used to precipitate DNA include ethanol and Isopropanol This process leaves a very pure sample for stringent DNA testing
47
PART B OBSERVING EXTRACTED DNA UNDER THE MICROSCOPE
48
PART B A CLOSER LOOK
8 Use a dropping pipette to carefully remove some of the threadlike substance from the top of your preparation
9 Place one or two drops onto the middle of a
microscope slide
10 Add two drops of methylene blue Wait 3 or 4 minutes to allow the methylene blue to be absorbed by the DNA
11 Carefully place a cover slip on the slide Gently press
a folded piece of paper towelling over the top of the prepared slide to soak up any excess liquid
12 Observe the DNA under low power then high power
49
DISCUSSIONANSWER IN GROUP RELATED WORKED 1 Write a detailed description of the material
floating at the top of the test tube after the alcohol was added
2 Describe the DNA as it appears under the
microscope under high power 3 Prepare a diagram of your DNA specimen 4 What was the reason for using methylene
blue
50
HOMEWORKCONSTRUCT A FLOW CHART DRAWING MIND MAP OF THE PROCESS YOU USED EXTRACTING THE DNA FROM KIWIrsquoS
Next to each step explain how the method you used was important (Hint What substances make up the membranes of cells and cell organelles
How do detergents work
What is the active ingredient in meat tenderiser
51
TIPS STEPS FOR FLOW CHART In order to release the DNA from the
nuclei of the pea cells you first separated the cells from one another
Then the cell and nuclear membranes needed to be ruptured to release the cell contents and the contents of the nucleus
Once removed from the nuclear membrane the DNA had to be untangled into the visible threadlike structures you ended up with
52
HOMEWORK EXTENSION GROUP QUESTIONS Where can DNA be found in the cell Discuss the action of the soap (detergent) on the cell
What is the purpose of the soap in this activity What was the purpose of the Sodium Chloride Include a
discussion of polarity and charged particles Why was the cold ethanol added to the soap and salt
mixture Describe the appearance of your final product Draw a diagram of DNA containing 5 sets of nucleotide
bases labelling the hydrogen bonds between the bases References and Resources Adapted from Berry Full
of DNA by Diane Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
53
HTTPWWWGENOMEBCCAEDUCATIONTEACHERSCLASSROOM-ACTIVITIESDNA-EXTRACTION
54
EXTRACHROMOSOMES EXIST IN PAIRS Because our chromosomes exist in pairs (and consequently we
have 2 alleles of each gene) we are a diploid species This is why our somatic cells are represented as 2n Our gametes (sperm and ova) on the other hand are haploid and are represented as n Other species may have different ploidy for example
triploid (3n) seedless watermelons tetaploid (4n) salmonidae fish pentaploid (5n) Kenai birch hexaploid (6n) some types of wheat kiwi fruit octaploid (8n) acipenser (a genus
of sturgeon fish strawberies) decaploid (10n) some strawberries dodecaploid (12n) some types of amphibians eg Xenopus
ruwenzoriensis
wwwcyberusca
55
YOUR EXTRACTED DNA UNDER THE MICROSCOPE LOOKED
LIKE
Collaboration-Brainstorming on what to look at under the
- DNA extraction- DNA is too small for even a microscope to see and after the extraction it just appears like a blob True or false
Could you any DNA strand
wwwemployeescsbsjuedu
56
HANDOUT ON DNA EXTRACTIONBACKGROUND EVERY DAY LIFE
EXTRA READING MATERIAL(EXTENSION )
DISCOVERING DNA DNA ON THE INSIDE USE OF DNA EXTRACTION INN EVERYDAY LIFE WHY IS DNA TESTING GOOD MOLECULAR BIOLOGY PCR-based diagnostics of genomic DNA
57
REFERENCES httpwwwsquidoocomhow-to-get-dna-from-a-kiwi-fruit Why Is DNA Testing Good | eHowcom
httpwwwehowcomabout_6684410_dna-testing-good_htmlixzz1MkbcIJs5 Why Is DNA Extraction Important | eHowcom
httpwwwehowcomlist_5839095_dna-extraction-important_htmlixzz1MkZDrqyY
Why Is Sodium Used in DNA Extraction | eHowcom httpwwwehowcomabout_6504902_sodium-used-dna-extraction_htmlixzz1MkaGWg57
Why Is Cold Alcohol Used in DNA Tests | eHowcom httpwwwehowcomabout_6399349_cold-alcohol-used-dna-tests_htmlixzz1MkbEmEdu
httplearngeneticsutaheducontentlabsextractionhowto www kamibudaksainsblogspotcom
httpwwwchemwatch goldcom References and Resources Adapted from Berry Full of DNA by Diane
Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
58
REFERENCES
Uses of DNA Extraction | eHowcom httpwwwehowcomabout_5344428_uses-dna-extractionhtmlixzz1MkWQB2TZ
Science 10 A Contextual Approach Heineman Queensland Science Projects Regan Spence Maggie Spenceley
httpenwikipediaorgwikiMolecular_biology httpwwwclinchemorgcgicontent
extract44102201 http
employeescsbsjueduhjakubowskiclassesch331dnaoldnastructurehtml
httpwwwgenomebccaeducationteachersclassroom-activitiesdna-extraction
26
INFORMATION DNA precipitates as a white stringy
ldquosnottyrdquo film at the water - alcohol interface and eventually will rise into the alcohol layer from the mixture layer
Allow test tube to sit for several minutes The clearer the DNA is the fewer
impurities you have
If you have an acceptable amount of DNA it can be spooled by rotating your collecting tool and then transferred into a clean tube container
27
WOW ndash SPOOLING THE DNA If you are careful you may be able wind up the DNA
around a glass rod or a skewer Position the tip of the glass rod or skewer where you can see the threads of DNA Steadily twist the rod or skewer as if you were making candy floss Alternatively use a straw to pull it out by suction ndash be careful not to get in you mouth
Donrsquot go too quickly
You should be able to pull the strands of DNA out of the mixture
ASK STUDENTS TO TAKE MOBILE PHONE PICTURES OF THEIR OWN EXTRACTED DNA AND COMPARE APPEARANCE WITH OTHERS
Photos of steps can be inserted in flow diagram(ICT)
28
KIWIDNA
29
If you want to save your DNA you can transfer it to a small container filled with alcohol
Leave tube container uncapped until the ethanol has evaporated
DNA can be stored in the fridge (dry or water buffer can be added)
You can now investigate the property
30
WHAT IS THAT STRINGY STUFFDNA IS A LONG STRINGY MOLECULE THE SALT THAT YOU ADDED IN STEP ONE HELPS IT STICK TOGETHER SO WHAT YOU SEE ARE CLUMPS OF TANGLED DNA MOLECULES
DNA NORMALLY STAYS DISSOLVED IN WATER BUT WHEN SALTY DNA COMES IN CONTACT WITH ALCOHOL IT BECOMES UNDISSOLVED THIS IS CALLED PRECIPITATION THE PHYSICAL FORCE OF THE DNA CLUMPING TOGETHER AS IT PRECIPITATES PULLS MORE STRANDS ALONG WITH IT AS IT RISES INTO THE ALCOHOL
31
THE WHYrsquoS Blending separated the pea cells In order to extract DNA from a cell the associated
membranes and proteins must first be removed (break apart the cells) and then physically separated (loosen the tough cell wall) from the DNA
To see the DNA we have to break open these two sacks We do this with detergent and salt( Sodium can be
involved in several of the steps ) Sodium is an element Its chemical symbol is Na for
Natrium the Latin word for sodium It is a positive ion and often associates with negative ions as part of useful compounds
Salt help precipitate protein and carbohydrates away from the DNA
Salt helps strip away the proteins associated with DNA + and ndash charge
32
THE WHYrsquoS Salt and Detergents are used to break down
cell walls and nuclear membranes to release the DNA
They work by chemically poking holes in the cell membranes or walls
Once holes are poked in the membranes the membranes can be further disrupted mechanically as with a blender
After that it is easier to get the contents of the cell out including the DNA
LETS HAVE A LOOK AT THE CELL
33
CELL TO DNA
httplearngeneticsutaheducontentlabsextractionhowtoHOW TO EXTRACT DNA FROM ANYTHING LIVING
34
THE WHYrsquoS WHY DETERGENT
Each cell is surrounded by a sack (the cell membrane)
DNA is found inside the second sack (nucleus) within each cell
To see the DNA we have to break it open A cell membrane has 2 layers of lipid(fat)
molecules with protein going through them When the lysis buffer (detergent) comes
close to the cell it captures the lipids and the proteins - breaks open the cell destroying the fatty membrane - DNA now released into the solution
35
A CELLS MEMBRANES HAVE TWO LAYERS OF LIPID (FAT) MOLECULES WITH PROTEINS GOING THROUGH THEM
36
Why detergent How does detergent work
Think about why you use soap to wash dishes or your hands To remove grease and dirt right
Soap molecules and grease molecules are made of two parts
1(Blue) Heads which like water 2(Green) Tails which hate water
37
AFTER ADDING THE DETERGENT WHAT DO YOU HAVE IN YOUR SOUP
38
THE WHYrsquoS ENZYME (MEAT TENDERIZER)
The DNA in the nucleus of the cell is molded folded and protected by proteins The tenderizer cut the proteins away from the DNA
In this experiment meat tenderizer acts as an enzyme to cut proteins just like a pair of scissors
The meat tenderizer cuts the proteins away from the DNA
39
THE WHYrsquoS CUTTING PROTEIN AND DNA
The DNA in the nucleus of the cell is moulded folded and protected by proteins
40
THE WHYrsquoSALCOHOL Alcohol is less dense than water so it floats
on top Look for clumps of white stringy stuff where the water and alcohol layers meet
DNA is not soluble in alcohol - other cell parts are
By adding alcohol DNA precipitates out of the solution and collect at the interface of the alcohol and soap layer
The colder the alcohol the less soluble the DNA will be in it
41
COLD ALCOHOL
DNA dissolves in water but precipitates in alcohol
Cold alcohol is used to separate DNA out of water-based solutions
This allows the DNA to be purified for subsequent genetic testing
Adding alcohol to a solution containing DNA is a simple way to obtain the pure DNA required and colder temperatures slow down enzymes that can break down DNA giving better extraction results
42
WHY IS DNA EXTRACTION IMPORTANT DNA extraction is an important molecular
biology procedure
By definition extraction is taking DNA out of any type of cell for the purpose of analysis
See Handouts for more information on the use of DNA extracted (extension only)
43
SUMMARIZEDNA EXTRACTING FROM KIWI FRUIT
44
BREAK DOWN CELL WALLS
Cells walls have to be destroyed to reach the DNA within cells
Extraction procedures to obtain pure DNA have to get rid of all the molecular and chemical components of the tissue from which the DNA is being extracted
First steps involve lysing or destroying the cell walls This can be done with a variety of chemical agents that are caustic to cell membranes but do not harm the DNA Cells can also be sonicated homogenized or ground up to destroy membranes
45
REMOVING Removing Lipids
Once the cell walls have been destroyed a detergent is added to get rid of the fats and oils that make up the cell membranes Detergents cause the fats and oils to dissolve into the solution
Remove ProteinsProteins and enzymes can be digested by
adding a protease to the solution Proteases break down proteins into small peptides and amino acids
46
PRECIPITATING AND PURIFYING Precipitate DNA
Adding cold alcohol to a solution will cause DNA to precipitate and aggregate The DNA can be collected by centrifuging the sample and pouring off the liquid layer The DNA should exist as a small pellet in the bottom of the centrifuge tube
Purify DNAThe DNA can be washed by re-suspending it
in a cold alcohol solution and re-centrifuging it several times to obtain a very pure DNA sample Typical alcohols used to precipitate DNA include ethanol and Isopropanol This process leaves a very pure sample for stringent DNA testing
47
PART B OBSERVING EXTRACTED DNA UNDER THE MICROSCOPE
48
PART B A CLOSER LOOK
8 Use a dropping pipette to carefully remove some of the threadlike substance from the top of your preparation
9 Place one or two drops onto the middle of a
microscope slide
10 Add two drops of methylene blue Wait 3 or 4 minutes to allow the methylene blue to be absorbed by the DNA
11 Carefully place a cover slip on the slide Gently press
a folded piece of paper towelling over the top of the prepared slide to soak up any excess liquid
12 Observe the DNA under low power then high power
49
DISCUSSIONANSWER IN GROUP RELATED WORKED 1 Write a detailed description of the material
floating at the top of the test tube after the alcohol was added
2 Describe the DNA as it appears under the
microscope under high power 3 Prepare a diagram of your DNA specimen 4 What was the reason for using methylene
blue
50
HOMEWORKCONSTRUCT A FLOW CHART DRAWING MIND MAP OF THE PROCESS YOU USED EXTRACTING THE DNA FROM KIWIrsquoS
Next to each step explain how the method you used was important (Hint What substances make up the membranes of cells and cell organelles
How do detergents work
What is the active ingredient in meat tenderiser
51
TIPS STEPS FOR FLOW CHART In order to release the DNA from the
nuclei of the pea cells you first separated the cells from one another
Then the cell and nuclear membranes needed to be ruptured to release the cell contents and the contents of the nucleus
Once removed from the nuclear membrane the DNA had to be untangled into the visible threadlike structures you ended up with
52
HOMEWORK EXTENSION GROUP QUESTIONS Where can DNA be found in the cell Discuss the action of the soap (detergent) on the cell
What is the purpose of the soap in this activity What was the purpose of the Sodium Chloride Include a
discussion of polarity and charged particles Why was the cold ethanol added to the soap and salt
mixture Describe the appearance of your final product Draw a diagram of DNA containing 5 sets of nucleotide
bases labelling the hydrogen bonds between the bases References and Resources Adapted from Berry Full
of DNA by Diane Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
53
HTTPWWWGENOMEBCCAEDUCATIONTEACHERSCLASSROOM-ACTIVITIESDNA-EXTRACTION
54
EXTRACHROMOSOMES EXIST IN PAIRS Because our chromosomes exist in pairs (and consequently we
have 2 alleles of each gene) we are a diploid species This is why our somatic cells are represented as 2n Our gametes (sperm and ova) on the other hand are haploid and are represented as n Other species may have different ploidy for example
triploid (3n) seedless watermelons tetaploid (4n) salmonidae fish pentaploid (5n) Kenai birch hexaploid (6n) some types of wheat kiwi fruit octaploid (8n) acipenser (a genus
of sturgeon fish strawberies) decaploid (10n) some strawberries dodecaploid (12n) some types of amphibians eg Xenopus
ruwenzoriensis
wwwcyberusca
55
YOUR EXTRACTED DNA UNDER THE MICROSCOPE LOOKED
LIKE
Collaboration-Brainstorming on what to look at under the
- DNA extraction- DNA is too small for even a microscope to see and after the extraction it just appears like a blob True or false
Could you any DNA strand
wwwemployeescsbsjuedu
56
HANDOUT ON DNA EXTRACTIONBACKGROUND EVERY DAY LIFE
EXTRA READING MATERIAL(EXTENSION )
DISCOVERING DNA DNA ON THE INSIDE USE OF DNA EXTRACTION INN EVERYDAY LIFE WHY IS DNA TESTING GOOD MOLECULAR BIOLOGY PCR-based diagnostics of genomic DNA
57
REFERENCES httpwwwsquidoocomhow-to-get-dna-from-a-kiwi-fruit Why Is DNA Testing Good | eHowcom
httpwwwehowcomabout_6684410_dna-testing-good_htmlixzz1MkbcIJs5 Why Is DNA Extraction Important | eHowcom
httpwwwehowcomlist_5839095_dna-extraction-important_htmlixzz1MkZDrqyY
Why Is Sodium Used in DNA Extraction | eHowcom httpwwwehowcomabout_6504902_sodium-used-dna-extraction_htmlixzz1MkaGWg57
Why Is Cold Alcohol Used in DNA Tests | eHowcom httpwwwehowcomabout_6399349_cold-alcohol-used-dna-tests_htmlixzz1MkbEmEdu
httplearngeneticsutaheducontentlabsextractionhowto www kamibudaksainsblogspotcom
httpwwwchemwatch goldcom References and Resources Adapted from Berry Full of DNA by Diane
Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
58
REFERENCES
Uses of DNA Extraction | eHowcom httpwwwehowcomabout_5344428_uses-dna-extractionhtmlixzz1MkWQB2TZ
Science 10 A Contextual Approach Heineman Queensland Science Projects Regan Spence Maggie Spenceley
httpenwikipediaorgwikiMolecular_biology httpwwwclinchemorgcgicontent
extract44102201 http
employeescsbsjueduhjakubowskiclassesch331dnaoldnastructurehtml
httpwwwgenomebccaeducationteachersclassroom-activitiesdna-extraction
27
WOW ndash SPOOLING THE DNA If you are careful you may be able wind up the DNA
around a glass rod or a skewer Position the tip of the glass rod or skewer where you can see the threads of DNA Steadily twist the rod or skewer as if you were making candy floss Alternatively use a straw to pull it out by suction ndash be careful not to get in you mouth
Donrsquot go too quickly
You should be able to pull the strands of DNA out of the mixture
ASK STUDENTS TO TAKE MOBILE PHONE PICTURES OF THEIR OWN EXTRACTED DNA AND COMPARE APPEARANCE WITH OTHERS
Photos of steps can be inserted in flow diagram(ICT)
28
KIWIDNA
29
If you want to save your DNA you can transfer it to a small container filled with alcohol
Leave tube container uncapped until the ethanol has evaporated
DNA can be stored in the fridge (dry or water buffer can be added)
You can now investigate the property
30
WHAT IS THAT STRINGY STUFFDNA IS A LONG STRINGY MOLECULE THE SALT THAT YOU ADDED IN STEP ONE HELPS IT STICK TOGETHER SO WHAT YOU SEE ARE CLUMPS OF TANGLED DNA MOLECULES
DNA NORMALLY STAYS DISSOLVED IN WATER BUT WHEN SALTY DNA COMES IN CONTACT WITH ALCOHOL IT BECOMES UNDISSOLVED THIS IS CALLED PRECIPITATION THE PHYSICAL FORCE OF THE DNA CLUMPING TOGETHER AS IT PRECIPITATES PULLS MORE STRANDS ALONG WITH IT AS IT RISES INTO THE ALCOHOL
31
THE WHYrsquoS Blending separated the pea cells In order to extract DNA from a cell the associated
membranes and proteins must first be removed (break apart the cells) and then physically separated (loosen the tough cell wall) from the DNA
To see the DNA we have to break open these two sacks We do this with detergent and salt( Sodium can be
involved in several of the steps ) Sodium is an element Its chemical symbol is Na for
Natrium the Latin word for sodium It is a positive ion and often associates with negative ions as part of useful compounds
Salt help precipitate protein and carbohydrates away from the DNA
Salt helps strip away the proteins associated with DNA + and ndash charge
32
THE WHYrsquoS Salt and Detergents are used to break down
cell walls and nuclear membranes to release the DNA
They work by chemically poking holes in the cell membranes or walls
Once holes are poked in the membranes the membranes can be further disrupted mechanically as with a blender
After that it is easier to get the contents of the cell out including the DNA
LETS HAVE A LOOK AT THE CELL
33
CELL TO DNA
httplearngeneticsutaheducontentlabsextractionhowtoHOW TO EXTRACT DNA FROM ANYTHING LIVING
34
THE WHYrsquoS WHY DETERGENT
Each cell is surrounded by a sack (the cell membrane)
DNA is found inside the second sack (nucleus) within each cell
To see the DNA we have to break it open A cell membrane has 2 layers of lipid(fat)
molecules with protein going through them When the lysis buffer (detergent) comes
close to the cell it captures the lipids and the proteins - breaks open the cell destroying the fatty membrane - DNA now released into the solution
35
A CELLS MEMBRANES HAVE TWO LAYERS OF LIPID (FAT) MOLECULES WITH PROTEINS GOING THROUGH THEM
36
Why detergent How does detergent work
Think about why you use soap to wash dishes or your hands To remove grease and dirt right
Soap molecules and grease molecules are made of two parts
1(Blue) Heads which like water 2(Green) Tails which hate water
37
AFTER ADDING THE DETERGENT WHAT DO YOU HAVE IN YOUR SOUP
38
THE WHYrsquoS ENZYME (MEAT TENDERIZER)
The DNA in the nucleus of the cell is molded folded and protected by proteins The tenderizer cut the proteins away from the DNA
In this experiment meat tenderizer acts as an enzyme to cut proteins just like a pair of scissors
The meat tenderizer cuts the proteins away from the DNA
39
THE WHYrsquoS CUTTING PROTEIN AND DNA
The DNA in the nucleus of the cell is moulded folded and protected by proteins
40
THE WHYrsquoSALCOHOL Alcohol is less dense than water so it floats
on top Look for clumps of white stringy stuff where the water and alcohol layers meet
DNA is not soluble in alcohol - other cell parts are
By adding alcohol DNA precipitates out of the solution and collect at the interface of the alcohol and soap layer
The colder the alcohol the less soluble the DNA will be in it
41
COLD ALCOHOL
DNA dissolves in water but precipitates in alcohol
Cold alcohol is used to separate DNA out of water-based solutions
This allows the DNA to be purified for subsequent genetic testing
Adding alcohol to a solution containing DNA is a simple way to obtain the pure DNA required and colder temperatures slow down enzymes that can break down DNA giving better extraction results
42
WHY IS DNA EXTRACTION IMPORTANT DNA extraction is an important molecular
biology procedure
By definition extraction is taking DNA out of any type of cell for the purpose of analysis
See Handouts for more information on the use of DNA extracted (extension only)
43
SUMMARIZEDNA EXTRACTING FROM KIWI FRUIT
44
BREAK DOWN CELL WALLS
Cells walls have to be destroyed to reach the DNA within cells
Extraction procedures to obtain pure DNA have to get rid of all the molecular and chemical components of the tissue from which the DNA is being extracted
First steps involve lysing or destroying the cell walls This can be done with a variety of chemical agents that are caustic to cell membranes but do not harm the DNA Cells can also be sonicated homogenized or ground up to destroy membranes
45
REMOVING Removing Lipids
Once the cell walls have been destroyed a detergent is added to get rid of the fats and oils that make up the cell membranes Detergents cause the fats and oils to dissolve into the solution
Remove ProteinsProteins and enzymes can be digested by
adding a protease to the solution Proteases break down proteins into small peptides and amino acids
46
PRECIPITATING AND PURIFYING Precipitate DNA
Adding cold alcohol to a solution will cause DNA to precipitate and aggregate The DNA can be collected by centrifuging the sample and pouring off the liquid layer The DNA should exist as a small pellet in the bottom of the centrifuge tube
Purify DNAThe DNA can be washed by re-suspending it
in a cold alcohol solution and re-centrifuging it several times to obtain a very pure DNA sample Typical alcohols used to precipitate DNA include ethanol and Isopropanol This process leaves a very pure sample for stringent DNA testing
47
PART B OBSERVING EXTRACTED DNA UNDER THE MICROSCOPE
48
PART B A CLOSER LOOK
8 Use a dropping pipette to carefully remove some of the threadlike substance from the top of your preparation
9 Place one or two drops onto the middle of a
microscope slide
10 Add two drops of methylene blue Wait 3 or 4 minutes to allow the methylene blue to be absorbed by the DNA
11 Carefully place a cover slip on the slide Gently press
a folded piece of paper towelling over the top of the prepared slide to soak up any excess liquid
12 Observe the DNA under low power then high power
49
DISCUSSIONANSWER IN GROUP RELATED WORKED 1 Write a detailed description of the material
floating at the top of the test tube after the alcohol was added
2 Describe the DNA as it appears under the
microscope under high power 3 Prepare a diagram of your DNA specimen 4 What was the reason for using methylene
blue
50
HOMEWORKCONSTRUCT A FLOW CHART DRAWING MIND MAP OF THE PROCESS YOU USED EXTRACTING THE DNA FROM KIWIrsquoS
Next to each step explain how the method you used was important (Hint What substances make up the membranes of cells and cell organelles
How do detergents work
What is the active ingredient in meat tenderiser
51
TIPS STEPS FOR FLOW CHART In order to release the DNA from the
nuclei of the pea cells you first separated the cells from one another
Then the cell and nuclear membranes needed to be ruptured to release the cell contents and the contents of the nucleus
Once removed from the nuclear membrane the DNA had to be untangled into the visible threadlike structures you ended up with
52
HOMEWORK EXTENSION GROUP QUESTIONS Where can DNA be found in the cell Discuss the action of the soap (detergent) on the cell
What is the purpose of the soap in this activity What was the purpose of the Sodium Chloride Include a
discussion of polarity and charged particles Why was the cold ethanol added to the soap and salt
mixture Describe the appearance of your final product Draw a diagram of DNA containing 5 sets of nucleotide
bases labelling the hydrogen bonds between the bases References and Resources Adapted from Berry Full
of DNA by Diane Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
53
HTTPWWWGENOMEBCCAEDUCATIONTEACHERSCLASSROOM-ACTIVITIESDNA-EXTRACTION
54
EXTRACHROMOSOMES EXIST IN PAIRS Because our chromosomes exist in pairs (and consequently we
have 2 alleles of each gene) we are a diploid species This is why our somatic cells are represented as 2n Our gametes (sperm and ova) on the other hand are haploid and are represented as n Other species may have different ploidy for example
triploid (3n) seedless watermelons tetaploid (4n) salmonidae fish pentaploid (5n) Kenai birch hexaploid (6n) some types of wheat kiwi fruit octaploid (8n) acipenser (a genus
of sturgeon fish strawberies) decaploid (10n) some strawberries dodecaploid (12n) some types of amphibians eg Xenopus
ruwenzoriensis
wwwcyberusca
55
YOUR EXTRACTED DNA UNDER THE MICROSCOPE LOOKED
LIKE
Collaboration-Brainstorming on what to look at under the
- DNA extraction- DNA is too small for even a microscope to see and after the extraction it just appears like a blob True or false
Could you any DNA strand
wwwemployeescsbsjuedu
56
HANDOUT ON DNA EXTRACTIONBACKGROUND EVERY DAY LIFE
EXTRA READING MATERIAL(EXTENSION )
DISCOVERING DNA DNA ON THE INSIDE USE OF DNA EXTRACTION INN EVERYDAY LIFE WHY IS DNA TESTING GOOD MOLECULAR BIOLOGY PCR-based diagnostics of genomic DNA
57
REFERENCES httpwwwsquidoocomhow-to-get-dna-from-a-kiwi-fruit Why Is DNA Testing Good | eHowcom
httpwwwehowcomabout_6684410_dna-testing-good_htmlixzz1MkbcIJs5 Why Is DNA Extraction Important | eHowcom
httpwwwehowcomlist_5839095_dna-extraction-important_htmlixzz1MkZDrqyY
Why Is Sodium Used in DNA Extraction | eHowcom httpwwwehowcomabout_6504902_sodium-used-dna-extraction_htmlixzz1MkaGWg57
Why Is Cold Alcohol Used in DNA Tests | eHowcom httpwwwehowcomabout_6399349_cold-alcohol-used-dna-tests_htmlixzz1MkbEmEdu
httplearngeneticsutaheducontentlabsextractionhowto www kamibudaksainsblogspotcom
httpwwwchemwatch goldcom References and Resources Adapted from Berry Full of DNA by Diane
Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
58
REFERENCES
Uses of DNA Extraction | eHowcom httpwwwehowcomabout_5344428_uses-dna-extractionhtmlixzz1MkWQB2TZ
Science 10 A Contextual Approach Heineman Queensland Science Projects Regan Spence Maggie Spenceley
httpenwikipediaorgwikiMolecular_biology httpwwwclinchemorgcgicontent
extract44102201 http
employeescsbsjueduhjakubowskiclassesch331dnaoldnastructurehtml
httpwwwgenomebccaeducationteachersclassroom-activitiesdna-extraction
28
KIWIDNA
29
If you want to save your DNA you can transfer it to a small container filled with alcohol
Leave tube container uncapped until the ethanol has evaporated
DNA can be stored in the fridge (dry or water buffer can be added)
You can now investigate the property
30
WHAT IS THAT STRINGY STUFFDNA IS A LONG STRINGY MOLECULE THE SALT THAT YOU ADDED IN STEP ONE HELPS IT STICK TOGETHER SO WHAT YOU SEE ARE CLUMPS OF TANGLED DNA MOLECULES
DNA NORMALLY STAYS DISSOLVED IN WATER BUT WHEN SALTY DNA COMES IN CONTACT WITH ALCOHOL IT BECOMES UNDISSOLVED THIS IS CALLED PRECIPITATION THE PHYSICAL FORCE OF THE DNA CLUMPING TOGETHER AS IT PRECIPITATES PULLS MORE STRANDS ALONG WITH IT AS IT RISES INTO THE ALCOHOL
31
THE WHYrsquoS Blending separated the pea cells In order to extract DNA from a cell the associated
membranes and proteins must first be removed (break apart the cells) and then physically separated (loosen the tough cell wall) from the DNA
To see the DNA we have to break open these two sacks We do this with detergent and salt( Sodium can be
involved in several of the steps ) Sodium is an element Its chemical symbol is Na for
Natrium the Latin word for sodium It is a positive ion and often associates with negative ions as part of useful compounds
Salt help precipitate protein and carbohydrates away from the DNA
Salt helps strip away the proteins associated with DNA + and ndash charge
32
THE WHYrsquoS Salt and Detergents are used to break down
cell walls and nuclear membranes to release the DNA
They work by chemically poking holes in the cell membranes or walls
Once holes are poked in the membranes the membranes can be further disrupted mechanically as with a blender
After that it is easier to get the contents of the cell out including the DNA
LETS HAVE A LOOK AT THE CELL
33
CELL TO DNA
httplearngeneticsutaheducontentlabsextractionhowtoHOW TO EXTRACT DNA FROM ANYTHING LIVING
34
THE WHYrsquoS WHY DETERGENT
Each cell is surrounded by a sack (the cell membrane)
DNA is found inside the second sack (nucleus) within each cell
To see the DNA we have to break it open A cell membrane has 2 layers of lipid(fat)
molecules with protein going through them When the lysis buffer (detergent) comes
close to the cell it captures the lipids and the proteins - breaks open the cell destroying the fatty membrane - DNA now released into the solution
35
A CELLS MEMBRANES HAVE TWO LAYERS OF LIPID (FAT) MOLECULES WITH PROTEINS GOING THROUGH THEM
36
Why detergent How does detergent work
Think about why you use soap to wash dishes or your hands To remove grease and dirt right
Soap molecules and grease molecules are made of two parts
1(Blue) Heads which like water 2(Green) Tails which hate water
37
AFTER ADDING THE DETERGENT WHAT DO YOU HAVE IN YOUR SOUP
38
THE WHYrsquoS ENZYME (MEAT TENDERIZER)
The DNA in the nucleus of the cell is molded folded and protected by proteins The tenderizer cut the proteins away from the DNA
In this experiment meat tenderizer acts as an enzyme to cut proteins just like a pair of scissors
The meat tenderizer cuts the proteins away from the DNA
39
THE WHYrsquoS CUTTING PROTEIN AND DNA
The DNA in the nucleus of the cell is moulded folded and protected by proteins
40
THE WHYrsquoSALCOHOL Alcohol is less dense than water so it floats
on top Look for clumps of white stringy stuff where the water and alcohol layers meet
DNA is not soluble in alcohol - other cell parts are
By adding alcohol DNA precipitates out of the solution and collect at the interface of the alcohol and soap layer
The colder the alcohol the less soluble the DNA will be in it
41
COLD ALCOHOL
DNA dissolves in water but precipitates in alcohol
Cold alcohol is used to separate DNA out of water-based solutions
This allows the DNA to be purified for subsequent genetic testing
Adding alcohol to a solution containing DNA is a simple way to obtain the pure DNA required and colder temperatures slow down enzymes that can break down DNA giving better extraction results
42
WHY IS DNA EXTRACTION IMPORTANT DNA extraction is an important molecular
biology procedure
By definition extraction is taking DNA out of any type of cell for the purpose of analysis
See Handouts for more information on the use of DNA extracted (extension only)
43
SUMMARIZEDNA EXTRACTING FROM KIWI FRUIT
44
BREAK DOWN CELL WALLS
Cells walls have to be destroyed to reach the DNA within cells
Extraction procedures to obtain pure DNA have to get rid of all the molecular and chemical components of the tissue from which the DNA is being extracted
First steps involve lysing or destroying the cell walls This can be done with a variety of chemical agents that are caustic to cell membranes but do not harm the DNA Cells can also be sonicated homogenized or ground up to destroy membranes
45
REMOVING Removing Lipids
Once the cell walls have been destroyed a detergent is added to get rid of the fats and oils that make up the cell membranes Detergents cause the fats and oils to dissolve into the solution
Remove ProteinsProteins and enzymes can be digested by
adding a protease to the solution Proteases break down proteins into small peptides and amino acids
46
PRECIPITATING AND PURIFYING Precipitate DNA
Adding cold alcohol to a solution will cause DNA to precipitate and aggregate The DNA can be collected by centrifuging the sample and pouring off the liquid layer The DNA should exist as a small pellet in the bottom of the centrifuge tube
Purify DNAThe DNA can be washed by re-suspending it
in a cold alcohol solution and re-centrifuging it several times to obtain a very pure DNA sample Typical alcohols used to precipitate DNA include ethanol and Isopropanol This process leaves a very pure sample for stringent DNA testing
47
PART B OBSERVING EXTRACTED DNA UNDER THE MICROSCOPE
48
PART B A CLOSER LOOK
8 Use a dropping pipette to carefully remove some of the threadlike substance from the top of your preparation
9 Place one or two drops onto the middle of a
microscope slide
10 Add two drops of methylene blue Wait 3 or 4 minutes to allow the methylene blue to be absorbed by the DNA
11 Carefully place a cover slip on the slide Gently press
a folded piece of paper towelling over the top of the prepared slide to soak up any excess liquid
12 Observe the DNA under low power then high power
49
DISCUSSIONANSWER IN GROUP RELATED WORKED 1 Write a detailed description of the material
floating at the top of the test tube after the alcohol was added
2 Describe the DNA as it appears under the
microscope under high power 3 Prepare a diagram of your DNA specimen 4 What was the reason for using methylene
blue
50
HOMEWORKCONSTRUCT A FLOW CHART DRAWING MIND MAP OF THE PROCESS YOU USED EXTRACTING THE DNA FROM KIWIrsquoS
Next to each step explain how the method you used was important (Hint What substances make up the membranes of cells and cell organelles
How do detergents work
What is the active ingredient in meat tenderiser
51
TIPS STEPS FOR FLOW CHART In order to release the DNA from the
nuclei of the pea cells you first separated the cells from one another
Then the cell and nuclear membranes needed to be ruptured to release the cell contents and the contents of the nucleus
Once removed from the nuclear membrane the DNA had to be untangled into the visible threadlike structures you ended up with
52
HOMEWORK EXTENSION GROUP QUESTIONS Where can DNA be found in the cell Discuss the action of the soap (detergent) on the cell
What is the purpose of the soap in this activity What was the purpose of the Sodium Chloride Include a
discussion of polarity and charged particles Why was the cold ethanol added to the soap and salt
mixture Describe the appearance of your final product Draw a diagram of DNA containing 5 sets of nucleotide
bases labelling the hydrogen bonds between the bases References and Resources Adapted from Berry Full
of DNA by Diane Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
53
HTTPWWWGENOMEBCCAEDUCATIONTEACHERSCLASSROOM-ACTIVITIESDNA-EXTRACTION
54
EXTRACHROMOSOMES EXIST IN PAIRS Because our chromosomes exist in pairs (and consequently we
have 2 alleles of each gene) we are a diploid species This is why our somatic cells are represented as 2n Our gametes (sperm and ova) on the other hand are haploid and are represented as n Other species may have different ploidy for example
triploid (3n) seedless watermelons tetaploid (4n) salmonidae fish pentaploid (5n) Kenai birch hexaploid (6n) some types of wheat kiwi fruit octaploid (8n) acipenser (a genus
of sturgeon fish strawberies) decaploid (10n) some strawberries dodecaploid (12n) some types of amphibians eg Xenopus
ruwenzoriensis
wwwcyberusca
55
YOUR EXTRACTED DNA UNDER THE MICROSCOPE LOOKED
LIKE
Collaboration-Brainstorming on what to look at under the
- DNA extraction- DNA is too small for even a microscope to see and after the extraction it just appears like a blob True or false
Could you any DNA strand
wwwemployeescsbsjuedu
56
HANDOUT ON DNA EXTRACTIONBACKGROUND EVERY DAY LIFE
EXTRA READING MATERIAL(EXTENSION )
DISCOVERING DNA DNA ON THE INSIDE USE OF DNA EXTRACTION INN EVERYDAY LIFE WHY IS DNA TESTING GOOD MOLECULAR BIOLOGY PCR-based diagnostics of genomic DNA
57
REFERENCES httpwwwsquidoocomhow-to-get-dna-from-a-kiwi-fruit Why Is DNA Testing Good | eHowcom
httpwwwehowcomabout_6684410_dna-testing-good_htmlixzz1MkbcIJs5 Why Is DNA Extraction Important | eHowcom
httpwwwehowcomlist_5839095_dna-extraction-important_htmlixzz1MkZDrqyY
Why Is Sodium Used in DNA Extraction | eHowcom httpwwwehowcomabout_6504902_sodium-used-dna-extraction_htmlixzz1MkaGWg57
Why Is Cold Alcohol Used in DNA Tests | eHowcom httpwwwehowcomabout_6399349_cold-alcohol-used-dna-tests_htmlixzz1MkbEmEdu
httplearngeneticsutaheducontentlabsextractionhowto www kamibudaksainsblogspotcom
httpwwwchemwatch goldcom References and Resources Adapted from Berry Full of DNA by Diane
Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
58
REFERENCES
Uses of DNA Extraction | eHowcom httpwwwehowcomabout_5344428_uses-dna-extractionhtmlixzz1MkWQB2TZ
Science 10 A Contextual Approach Heineman Queensland Science Projects Regan Spence Maggie Spenceley
httpenwikipediaorgwikiMolecular_biology httpwwwclinchemorgcgicontent
extract44102201 http
employeescsbsjueduhjakubowskiclassesch331dnaoldnastructurehtml
httpwwwgenomebccaeducationteachersclassroom-activitiesdna-extraction
29
If you want to save your DNA you can transfer it to a small container filled with alcohol
Leave tube container uncapped until the ethanol has evaporated
DNA can be stored in the fridge (dry or water buffer can be added)
You can now investigate the property
30
WHAT IS THAT STRINGY STUFFDNA IS A LONG STRINGY MOLECULE THE SALT THAT YOU ADDED IN STEP ONE HELPS IT STICK TOGETHER SO WHAT YOU SEE ARE CLUMPS OF TANGLED DNA MOLECULES
DNA NORMALLY STAYS DISSOLVED IN WATER BUT WHEN SALTY DNA COMES IN CONTACT WITH ALCOHOL IT BECOMES UNDISSOLVED THIS IS CALLED PRECIPITATION THE PHYSICAL FORCE OF THE DNA CLUMPING TOGETHER AS IT PRECIPITATES PULLS MORE STRANDS ALONG WITH IT AS IT RISES INTO THE ALCOHOL
31
THE WHYrsquoS Blending separated the pea cells In order to extract DNA from a cell the associated
membranes and proteins must first be removed (break apart the cells) and then physically separated (loosen the tough cell wall) from the DNA
To see the DNA we have to break open these two sacks We do this with detergent and salt( Sodium can be
involved in several of the steps ) Sodium is an element Its chemical symbol is Na for
Natrium the Latin word for sodium It is a positive ion and often associates with negative ions as part of useful compounds
Salt help precipitate protein and carbohydrates away from the DNA
Salt helps strip away the proteins associated with DNA + and ndash charge
32
THE WHYrsquoS Salt and Detergents are used to break down
cell walls and nuclear membranes to release the DNA
They work by chemically poking holes in the cell membranes or walls
Once holes are poked in the membranes the membranes can be further disrupted mechanically as with a blender
After that it is easier to get the contents of the cell out including the DNA
LETS HAVE A LOOK AT THE CELL
33
CELL TO DNA
httplearngeneticsutaheducontentlabsextractionhowtoHOW TO EXTRACT DNA FROM ANYTHING LIVING
34
THE WHYrsquoS WHY DETERGENT
Each cell is surrounded by a sack (the cell membrane)
DNA is found inside the second sack (nucleus) within each cell
To see the DNA we have to break it open A cell membrane has 2 layers of lipid(fat)
molecules with protein going through them When the lysis buffer (detergent) comes
close to the cell it captures the lipids and the proteins - breaks open the cell destroying the fatty membrane - DNA now released into the solution
35
A CELLS MEMBRANES HAVE TWO LAYERS OF LIPID (FAT) MOLECULES WITH PROTEINS GOING THROUGH THEM
36
Why detergent How does detergent work
Think about why you use soap to wash dishes or your hands To remove grease and dirt right
Soap molecules and grease molecules are made of two parts
1(Blue) Heads which like water 2(Green) Tails which hate water
37
AFTER ADDING THE DETERGENT WHAT DO YOU HAVE IN YOUR SOUP
38
THE WHYrsquoS ENZYME (MEAT TENDERIZER)
The DNA in the nucleus of the cell is molded folded and protected by proteins The tenderizer cut the proteins away from the DNA
In this experiment meat tenderizer acts as an enzyme to cut proteins just like a pair of scissors
The meat tenderizer cuts the proteins away from the DNA
39
THE WHYrsquoS CUTTING PROTEIN AND DNA
The DNA in the nucleus of the cell is moulded folded and protected by proteins
40
THE WHYrsquoSALCOHOL Alcohol is less dense than water so it floats
on top Look for clumps of white stringy stuff where the water and alcohol layers meet
DNA is not soluble in alcohol - other cell parts are
By adding alcohol DNA precipitates out of the solution and collect at the interface of the alcohol and soap layer
The colder the alcohol the less soluble the DNA will be in it
41
COLD ALCOHOL
DNA dissolves in water but precipitates in alcohol
Cold alcohol is used to separate DNA out of water-based solutions
This allows the DNA to be purified for subsequent genetic testing
Adding alcohol to a solution containing DNA is a simple way to obtain the pure DNA required and colder temperatures slow down enzymes that can break down DNA giving better extraction results
42
WHY IS DNA EXTRACTION IMPORTANT DNA extraction is an important molecular
biology procedure
By definition extraction is taking DNA out of any type of cell for the purpose of analysis
See Handouts for more information on the use of DNA extracted (extension only)
43
SUMMARIZEDNA EXTRACTING FROM KIWI FRUIT
44
BREAK DOWN CELL WALLS
Cells walls have to be destroyed to reach the DNA within cells
Extraction procedures to obtain pure DNA have to get rid of all the molecular and chemical components of the tissue from which the DNA is being extracted
First steps involve lysing or destroying the cell walls This can be done with a variety of chemical agents that are caustic to cell membranes but do not harm the DNA Cells can also be sonicated homogenized or ground up to destroy membranes
45
REMOVING Removing Lipids
Once the cell walls have been destroyed a detergent is added to get rid of the fats and oils that make up the cell membranes Detergents cause the fats and oils to dissolve into the solution
Remove ProteinsProteins and enzymes can be digested by
adding a protease to the solution Proteases break down proteins into small peptides and amino acids
46
PRECIPITATING AND PURIFYING Precipitate DNA
Adding cold alcohol to a solution will cause DNA to precipitate and aggregate The DNA can be collected by centrifuging the sample and pouring off the liquid layer The DNA should exist as a small pellet in the bottom of the centrifuge tube
Purify DNAThe DNA can be washed by re-suspending it
in a cold alcohol solution and re-centrifuging it several times to obtain a very pure DNA sample Typical alcohols used to precipitate DNA include ethanol and Isopropanol This process leaves a very pure sample for stringent DNA testing
47
PART B OBSERVING EXTRACTED DNA UNDER THE MICROSCOPE
48
PART B A CLOSER LOOK
8 Use a dropping pipette to carefully remove some of the threadlike substance from the top of your preparation
9 Place one or two drops onto the middle of a
microscope slide
10 Add two drops of methylene blue Wait 3 or 4 minutes to allow the methylene blue to be absorbed by the DNA
11 Carefully place a cover slip on the slide Gently press
a folded piece of paper towelling over the top of the prepared slide to soak up any excess liquid
12 Observe the DNA under low power then high power
49
DISCUSSIONANSWER IN GROUP RELATED WORKED 1 Write a detailed description of the material
floating at the top of the test tube after the alcohol was added
2 Describe the DNA as it appears under the
microscope under high power 3 Prepare a diagram of your DNA specimen 4 What was the reason for using methylene
blue
50
HOMEWORKCONSTRUCT A FLOW CHART DRAWING MIND MAP OF THE PROCESS YOU USED EXTRACTING THE DNA FROM KIWIrsquoS
Next to each step explain how the method you used was important (Hint What substances make up the membranes of cells and cell organelles
How do detergents work
What is the active ingredient in meat tenderiser
51
TIPS STEPS FOR FLOW CHART In order to release the DNA from the
nuclei of the pea cells you first separated the cells from one another
Then the cell and nuclear membranes needed to be ruptured to release the cell contents and the contents of the nucleus
Once removed from the nuclear membrane the DNA had to be untangled into the visible threadlike structures you ended up with
52
HOMEWORK EXTENSION GROUP QUESTIONS Where can DNA be found in the cell Discuss the action of the soap (detergent) on the cell
What is the purpose of the soap in this activity What was the purpose of the Sodium Chloride Include a
discussion of polarity and charged particles Why was the cold ethanol added to the soap and salt
mixture Describe the appearance of your final product Draw a diagram of DNA containing 5 sets of nucleotide
bases labelling the hydrogen bonds between the bases References and Resources Adapted from Berry Full
of DNA by Diane Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
53
HTTPWWWGENOMEBCCAEDUCATIONTEACHERSCLASSROOM-ACTIVITIESDNA-EXTRACTION
54
EXTRACHROMOSOMES EXIST IN PAIRS Because our chromosomes exist in pairs (and consequently we
have 2 alleles of each gene) we are a diploid species This is why our somatic cells are represented as 2n Our gametes (sperm and ova) on the other hand are haploid and are represented as n Other species may have different ploidy for example
triploid (3n) seedless watermelons tetaploid (4n) salmonidae fish pentaploid (5n) Kenai birch hexaploid (6n) some types of wheat kiwi fruit octaploid (8n) acipenser (a genus
of sturgeon fish strawberies) decaploid (10n) some strawberries dodecaploid (12n) some types of amphibians eg Xenopus
ruwenzoriensis
wwwcyberusca
55
YOUR EXTRACTED DNA UNDER THE MICROSCOPE LOOKED
LIKE
Collaboration-Brainstorming on what to look at under the
- DNA extraction- DNA is too small for even a microscope to see and after the extraction it just appears like a blob True or false
Could you any DNA strand
wwwemployeescsbsjuedu
56
HANDOUT ON DNA EXTRACTIONBACKGROUND EVERY DAY LIFE
EXTRA READING MATERIAL(EXTENSION )
DISCOVERING DNA DNA ON THE INSIDE USE OF DNA EXTRACTION INN EVERYDAY LIFE WHY IS DNA TESTING GOOD MOLECULAR BIOLOGY PCR-based diagnostics of genomic DNA
57
REFERENCES httpwwwsquidoocomhow-to-get-dna-from-a-kiwi-fruit Why Is DNA Testing Good | eHowcom
httpwwwehowcomabout_6684410_dna-testing-good_htmlixzz1MkbcIJs5 Why Is DNA Extraction Important | eHowcom
httpwwwehowcomlist_5839095_dna-extraction-important_htmlixzz1MkZDrqyY
Why Is Sodium Used in DNA Extraction | eHowcom httpwwwehowcomabout_6504902_sodium-used-dna-extraction_htmlixzz1MkaGWg57
Why Is Cold Alcohol Used in DNA Tests | eHowcom httpwwwehowcomabout_6399349_cold-alcohol-used-dna-tests_htmlixzz1MkbEmEdu
httplearngeneticsutaheducontentlabsextractionhowto www kamibudaksainsblogspotcom
httpwwwchemwatch goldcom References and Resources Adapted from Berry Full of DNA by Diane
Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
58
REFERENCES
Uses of DNA Extraction | eHowcom httpwwwehowcomabout_5344428_uses-dna-extractionhtmlixzz1MkWQB2TZ
Science 10 A Contextual Approach Heineman Queensland Science Projects Regan Spence Maggie Spenceley
httpenwikipediaorgwikiMolecular_biology httpwwwclinchemorgcgicontent
extract44102201 http
employeescsbsjueduhjakubowskiclassesch331dnaoldnastructurehtml
httpwwwgenomebccaeducationteachersclassroom-activitiesdna-extraction
30
WHAT IS THAT STRINGY STUFFDNA IS A LONG STRINGY MOLECULE THE SALT THAT YOU ADDED IN STEP ONE HELPS IT STICK TOGETHER SO WHAT YOU SEE ARE CLUMPS OF TANGLED DNA MOLECULES
DNA NORMALLY STAYS DISSOLVED IN WATER BUT WHEN SALTY DNA COMES IN CONTACT WITH ALCOHOL IT BECOMES UNDISSOLVED THIS IS CALLED PRECIPITATION THE PHYSICAL FORCE OF THE DNA CLUMPING TOGETHER AS IT PRECIPITATES PULLS MORE STRANDS ALONG WITH IT AS IT RISES INTO THE ALCOHOL
31
THE WHYrsquoS Blending separated the pea cells In order to extract DNA from a cell the associated
membranes and proteins must first be removed (break apart the cells) and then physically separated (loosen the tough cell wall) from the DNA
To see the DNA we have to break open these two sacks We do this with detergent and salt( Sodium can be
involved in several of the steps ) Sodium is an element Its chemical symbol is Na for
Natrium the Latin word for sodium It is a positive ion and often associates with negative ions as part of useful compounds
Salt help precipitate protein and carbohydrates away from the DNA
Salt helps strip away the proteins associated with DNA + and ndash charge
32
THE WHYrsquoS Salt and Detergents are used to break down
cell walls and nuclear membranes to release the DNA
They work by chemically poking holes in the cell membranes or walls
Once holes are poked in the membranes the membranes can be further disrupted mechanically as with a blender
After that it is easier to get the contents of the cell out including the DNA
LETS HAVE A LOOK AT THE CELL
33
CELL TO DNA
httplearngeneticsutaheducontentlabsextractionhowtoHOW TO EXTRACT DNA FROM ANYTHING LIVING
34
THE WHYrsquoS WHY DETERGENT
Each cell is surrounded by a sack (the cell membrane)
DNA is found inside the second sack (nucleus) within each cell
To see the DNA we have to break it open A cell membrane has 2 layers of lipid(fat)
molecules with protein going through them When the lysis buffer (detergent) comes
close to the cell it captures the lipids and the proteins - breaks open the cell destroying the fatty membrane - DNA now released into the solution
35
A CELLS MEMBRANES HAVE TWO LAYERS OF LIPID (FAT) MOLECULES WITH PROTEINS GOING THROUGH THEM
36
Why detergent How does detergent work
Think about why you use soap to wash dishes or your hands To remove grease and dirt right
Soap molecules and grease molecules are made of two parts
1(Blue) Heads which like water 2(Green) Tails which hate water
37
AFTER ADDING THE DETERGENT WHAT DO YOU HAVE IN YOUR SOUP
38
THE WHYrsquoS ENZYME (MEAT TENDERIZER)
The DNA in the nucleus of the cell is molded folded and protected by proteins The tenderizer cut the proteins away from the DNA
In this experiment meat tenderizer acts as an enzyme to cut proteins just like a pair of scissors
The meat tenderizer cuts the proteins away from the DNA
39
THE WHYrsquoS CUTTING PROTEIN AND DNA
The DNA in the nucleus of the cell is moulded folded and protected by proteins
40
THE WHYrsquoSALCOHOL Alcohol is less dense than water so it floats
on top Look for clumps of white stringy stuff where the water and alcohol layers meet
DNA is not soluble in alcohol - other cell parts are
By adding alcohol DNA precipitates out of the solution and collect at the interface of the alcohol and soap layer
The colder the alcohol the less soluble the DNA will be in it
41
COLD ALCOHOL
DNA dissolves in water but precipitates in alcohol
Cold alcohol is used to separate DNA out of water-based solutions
This allows the DNA to be purified for subsequent genetic testing
Adding alcohol to a solution containing DNA is a simple way to obtain the pure DNA required and colder temperatures slow down enzymes that can break down DNA giving better extraction results
42
WHY IS DNA EXTRACTION IMPORTANT DNA extraction is an important molecular
biology procedure
By definition extraction is taking DNA out of any type of cell for the purpose of analysis
See Handouts for more information on the use of DNA extracted (extension only)
43
SUMMARIZEDNA EXTRACTING FROM KIWI FRUIT
44
BREAK DOWN CELL WALLS
Cells walls have to be destroyed to reach the DNA within cells
Extraction procedures to obtain pure DNA have to get rid of all the molecular and chemical components of the tissue from which the DNA is being extracted
First steps involve lysing or destroying the cell walls This can be done with a variety of chemical agents that are caustic to cell membranes but do not harm the DNA Cells can also be sonicated homogenized or ground up to destroy membranes
45
REMOVING Removing Lipids
Once the cell walls have been destroyed a detergent is added to get rid of the fats and oils that make up the cell membranes Detergents cause the fats and oils to dissolve into the solution
Remove ProteinsProteins and enzymes can be digested by
adding a protease to the solution Proteases break down proteins into small peptides and amino acids
46
PRECIPITATING AND PURIFYING Precipitate DNA
Adding cold alcohol to a solution will cause DNA to precipitate and aggregate The DNA can be collected by centrifuging the sample and pouring off the liquid layer The DNA should exist as a small pellet in the bottom of the centrifuge tube
Purify DNAThe DNA can be washed by re-suspending it
in a cold alcohol solution and re-centrifuging it several times to obtain a very pure DNA sample Typical alcohols used to precipitate DNA include ethanol and Isopropanol This process leaves a very pure sample for stringent DNA testing
47
PART B OBSERVING EXTRACTED DNA UNDER THE MICROSCOPE
48
PART B A CLOSER LOOK
8 Use a dropping pipette to carefully remove some of the threadlike substance from the top of your preparation
9 Place one or two drops onto the middle of a
microscope slide
10 Add two drops of methylene blue Wait 3 or 4 minutes to allow the methylene blue to be absorbed by the DNA
11 Carefully place a cover slip on the slide Gently press
a folded piece of paper towelling over the top of the prepared slide to soak up any excess liquid
12 Observe the DNA under low power then high power
49
DISCUSSIONANSWER IN GROUP RELATED WORKED 1 Write a detailed description of the material
floating at the top of the test tube after the alcohol was added
2 Describe the DNA as it appears under the
microscope under high power 3 Prepare a diagram of your DNA specimen 4 What was the reason for using methylene
blue
50
HOMEWORKCONSTRUCT A FLOW CHART DRAWING MIND MAP OF THE PROCESS YOU USED EXTRACTING THE DNA FROM KIWIrsquoS
Next to each step explain how the method you used was important (Hint What substances make up the membranes of cells and cell organelles
How do detergents work
What is the active ingredient in meat tenderiser
51
TIPS STEPS FOR FLOW CHART In order to release the DNA from the
nuclei of the pea cells you first separated the cells from one another
Then the cell and nuclear membranes needed to be ruptured to release the cell contents and the contents of the nucleus
Once removed from the nuclear membrane the DNA had to be untangled into the visible threadlike structures you ended up with
52
HOMEWORK EXTENSION GROUP QUESTIONS Where can DNA be found in the cell Discuss the action of the soap (detergent) on the cell
What is the purpose of the soap in this activity What was the purpose of the Sodium Chloride Include a
discussion of polarity and charged particles Why was the cold ethanol added to the soap and salt
mixture Describe the appearance of your final product Draw a diagram of DNA containing 5 sets of nucleotide
bases labelling the hydrogen bonds between the bases References and Resources Adapted from Berry Full
of DNA by Diane Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
53
HTTPWWWGENOMEBCCAEDUCATIONTEACHERSCLASSROOM-ACTIVITIESDNA-EXTRACTION
54
EXTRACHROMOSOMES EXIST IN PAIRS Because our chromosomes exist in pairs (and consequently we
have 2 alleles of each gene) we are a diploid species This is why our somatic cells are represented as 2n Our gametes (sperm and ova) on the other hand are haploid and are represented as n Other species may have different ploidy for example
triploid (3n) seedless watermelons tetaploid (4n) salmonidae fish pentaploid (5n) Kenai birch hexaploid (6n) some types of wheat kiwi fruit octaploid (8n) acipenser (a genus
of sturgeon fish strawberies) decaploid (10n) some strawberries dodecaploid (12n) some types of amphibians eg Xenopus
ruwenzoriensis
wwwcyberusca
55
YOUR EXTRACTED DNA UNDER THE MICROSCOPE LOOKED
LIKE
Collaboration-Brainstorming on what to look at under the
- DNA extraction- DNA is too small for even a microscope to see and after the extraction it just appears like a blob True or false
Could you any DNA strand
wwwemployeescsbsjuedu
56
HANDOUT ON DNA EXTRACTIONBACKGROUND EVERY DAY LIFE
EXTRA READING MATERIAL(EXTENSION )
DISCOVERING DNA DNA ON THE INSIDE USE OF DNA EXTRACTION INN EVERYDAY LIFE WHY IS DNA TESTING GOOD MOLECULAR BIOLOGY PCR-based diagnostics of genomic DNA
57
REFERENCES httpwwwsquidoocomhow-to-get-dna-from-a-kiwi-fruit Why Is DNA Testing Good | eHowcom
httpwwwehowcomabout_6684410_dna-testing-good_htmlixzz1MkbcIJs5 Why Is DNA Extraction Important | eHowcom
httpwwwehowcomlist_5839095_dna-extraction-important_htmlixzz1MkZDrqyY
Why Is Sodium Used in DNA Extraction | eHowcom httpwwwehowcomabout_6504902_sodium-used-dna-extraction_htmlixzz1MkaGWg57
Why Is Cold Alcohol Used in DNA Tests | eHowcom httpwwwehowcomabout_6399349_cold-alcohol-used-dna-tests_htmlixzz1MkbEmEdu
httplearngeneticsutaheducontentlabsextractionhowto www kamibudaksainsblogspotcom
httpwwwchemwatch goldcom References and Resources Adapted from Berry Full of DNA by Diane
Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
58
REFERENCES
Uses of DNA Extraction | eHowcom httpwwwehowcomabout_5344428_uses-dna-extractionhtmlixzz1MkWQB2TZ
Science 10 A Contextual Approach Heineman Queensland Science Projects Regan Spence Maggie Spenceley
httpenwikipediaorgwikiMolecular_biology httpwwwclinchemorgcgicontent
extract44102201 http
employeescsbsjueduhjakubowskiclassesch331dnaoldnastructurehtml
httpwwwgenomebccaeducationteachersclassroom-activitiesdna-extraction
31
THE WHYrsquoS Blending separated the pea cells In order to extract DNA from a cell the associated
membranes and proteins must first be removed (break apart the cells) and then physically separated (loosen the tough cell wall) from the DNA
To see the DNA we have to break open these two sacks We do this with detergent and salt( Sodium can be
involved in several of the steps ) Sodium is an element Its chemical symbol is Na for
Natrium the Latin word for sodium It is a positive ion and often associates with negative ions as part of useful compounds
Salt help precipitate protein and carbohydrates away from the DNA
Salt helps strip away the proteins associated with DNA + and ndash charge
32
THE WHYrsquoS Salt and Detergents are used to break down
cell walls and nuclear membranes to release the DNA
They work by chemically poking holes in the cell membranes or walls
Once holes are poked in the membranes the membranes can be further disrupted mechanically as with a blender
After that it is easier to get the contents of the cell out including the DNA
LETS HAVE A LOOK AT THE CELL
33
CELL TO DNA
httplearngeneticsutaheducontentlabsextractionhowtoHOW TO EXTRACT DNA FROM ANYTHING LIVING
34
THE WHYrsquoS WHY DETERGENT
Each cell is surrounded by a sack (the cell membrane)
DNA is found inside the second sack (nucleus) within each cell
To see the DNA we have to break it open A cell membrane has 2 layers of lipid(fat)
molecules with protein going through them When the lysis buffer (detergent) comes
close to the cell it captures the lipids and the proteins - breaks open the cell destroying the fatty membrane - DNA now released into the solution
35
A CELLS MEMBRANES HAVE TWO LAYERS OF LIPID (FAT) MOLECULES WITH PROTEINS GOING THROUGH THEM
36
Why detergent How does detergent work
Think about why you use soap to wash dishes or your hands To remove grease and dirt right
Soap molecules and grease molecules are made of two parts
1(Blue) Heads which like water 2(Green) Tails which hate water
37
AFTER ADDING THE DETERGENT WHAT DO YOU HAVE IN YOUR SOUP
38
THE WHYrsquoS ENZYME (MEAT TENDERIZER)
The DNA in the nucleus of the cell is molded folded and protected by proteins The tenderizer cut the proteins away from the DNA
In this experiment meat tenderizer acts as an enzyme to cut proteins just like a pair of scissors
The meat tenderizer cuts the proteins away from the DNA
39
THE WHYrsquoS CUTTING PROTEIN AND DNA
The DNA in the nucleus of the cell is moulded folded and protected by proteins
40
THE WHYrsquoSALCOHOL Alcohol is less dense than water so it floats
on top Look for clumps of white stringy stuff where the water and alcohol layers meet
DNA is not soluble in alcohol - other cell parts are
By adding alcohol DNA precipitates out of the solution and collect at the interface of the alcohol and soap layer
The colder the alcohol the less soluble the DNA will be in it
41
COLD ALCOHOL
DNA dissolves in water but precipitates in alcohol
Cold alcohol is used to separate DNA out of water-based solutions
This allows the DNA to be purified for subsequent genetic testing
Adding alcohol to a solution containing DNA is a simple way to obtain the pure DNA required and colder temperatures slow down enzymes that can break down DNA giving better extraction results
42
WHY IS DNA EXTRACTION IMPORTANT DNA extraction is an important molecular
biology procedure
By definition extraction is taking DNA out of any type of cell for the purpose of analysis
See Handouts for more information on the use of DNA extracted (extension only)
43
SUMMARIZEDNA EXTRACTING FROM KIWI FRUIT
44
BREAK DOWN CELL WALLS
Cells walls have to be destroyed to reach the DNA within cells
Extraction procedures to obtain pure DNA have to get rid of all the molecular and chemical components of the tissue from which the DNA is being extracted
First steps involve lysing or destroying the cell walls This can be done with a variety of chemical agents that are caustic to cell membranes but do not harm the DNA Cells can also be sonicated homogenized or ground up to destroy membranes
45
REMOVING Removing Lipids
Once the cell walls have been destroyed a detergent is added to get rid of the fats and oils that make up the cell membranes Detergents cause the fats and oils to dissolve into the solution
Remove ProteinsProteins and enzymes can be digested by
adding a protease to the solution Proteases break down proteins into small peptides and amino acids
46
PRECIPITATING AND PURIFYING Precipitate DNA
Adding cold alcohol to a solution will cause DNA to precipitate and aggregate The DNA can be collected by centrifuging the sample and pouring off the liquid layer The DNA should exist as a small pellet in the bottom of the centrifuge tube
Purify DNAThe DNA can be washed by re-suspending it
in a cold alcohol solution and re-centrifuging it several times to obtain a very pure DNA sample Typical alcohols used to precipitate DNA include ethanol and Isopropanol This process leaves a very pure sample for stringent DNA testing
47
PART B OBSERVING EXTRACTED DNA UNDER THE MICROSCOPE
48
PART B A CLOSER LOOK
8 Use a dropping pipette to carefully remove some of the threadlike substance from the top of your preparation
9 Place one or two drops onto the middle of a
microscope slide
10 Add two drops of methylene blue Wait 3 or 4 minutes to allow the methylene blue to be absorbed by the DNA
11 Carefully place a cover slip on the slide Gently press
a folded piece of paper towelling over the top of the prepared slide to soak up any excess liquid
12 Observe the DNA under low power then high power
49
DISCUSSIONANSWER IN GROUP RELATED WORKED 1 Write a detailed description of the material
floating at the top of the test tube after the alcohol was added
2 Describe the DNA as it appears under the
microscope under high power 3 Prepare a diagram of your DNA specimen 4 What was the reason for using methylene
blue
50
HOMEWORKCONSTRUCT A FLOW CHART DRAWING MIND MAP OF THE PROCESS YOU USED EXTRACTING THE DNA FROM KIWIrsquoS
Next to each step explain how the method you used was important (Hint What substances make up the membranes of cells and cell organelles
How do detergents work
What is the active ingredient in meat tenderiser
51
TIPS STEPS FOR FLOW CHART In order to release the DNA from the
nuclei of the pea cells you first separated the cells from one another
Then the cell and nuclear membranes needed to be ruptured to release the cell contents and the contents of the nucleus
Once removed from the nuclear membrane the DNA had to be untangled into the visible threadlike structures you ended up with
52
HOMEWORK EXTENSION GROUP QUESTIONS Where can DNA be found in the cell Discuss the action of the soap (detergent) on the cell
What is the purpose of the soap in this activity What was the purpose of the Sodium Chloride Include a
discussion of polarity and charged particles Why was the cold ethanol added to the soap and salt
mixture Describe the appearance of your final product Draw a diagram of DNA containing 5 sets of nucleotide
bases labelling the hydrogen bonds between the bases References and Resources Adapted from Berry Full
of DNA by Diane Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
53
HTTPWWWGENOMEBCCAEDUCATIONTEACHERSCLASSROOM-ACTIVITIESDNA-EXTRACTION
54
EXTRACHROMOSOMES EXIST IN PAIRS Because our chromosomes exist in pairs (and consequently we
have 2 alleles of each gene) we are a diploid species This is why our somatic cells are represented as 2n Our gametes (sperm and ova) on the other hand are haploid and are represented as n Other species may have different ploidy for example
triploid (3n) seedless watermelons tetaploid (4n) salmonidae fish pentaploid (5n) Kenai birch hexaploid (6n) some types of wheat kiwi fruit octaploid (8n) acipenser (a genus
of sturgeon fish strawberies) decaploid (10n) some strawberries dodecaploid (12n) some types of amphibians eg Xenopus
ruwenzoriensis
wwwcyberusca
55
YOUR EXTRACTED DNA UNDER THE MICROSCOPE LOOKED
LIKE
Collaboration-Brainstorming on what to look at under the
- DNA extraction- DNA is too small for even a microscope to see and after the extraction it just appears like a blob True or false
Could you any DNA strand
wwwemployeescsbsjuedu
56
HANDOUT ON DNA EXTRACTIONBACKGROUND EVERY DAY LIFE
EXTRA READING MATERIAL(EXTENSION )
DISCOVERING DNA DNA ON THE INSIDE USE OF DNA EXTRACTION INN EVERYDAY LIFE WHY IS DNA TESTING GOOD MOLECULAR BIOLOGY PCR-based diagnostics of genomic DNA
57
REFERENCES httpwwwsquidoocomhow-to-get-dna-from-a-kiwi-fruit Why Is DNA Testing Good | eHowcom
httpwwwehowcomabout_6684410_dna-testing-good_htmlixzz1MkbcIJs5 Why Is DNA Extraction Important | eHowcom
httpwwwehowcomlist_5839095_dna-extraction-important_htmlixzz1MkZDrqyY
Why Is Sodium Used in DNA Extraction | eHowcom httpwwwehowcomabout_6504902_sodium-used-dna-extraction_htmlixzz1MkaGWg57
Why Is Cold Alcohol Used in DNA Tests | eHowcom httpwwwehowcomabout_6399349_cold-alcohol-used-dna-tests_htmlixzz1MkbEmEdu
httplearngeneticsutaheducontentlabsextractionhowto www kamibudaksainsblogspotcom
httpwwwchemwatch goldcom References and Resources Adapted from Berry Full of DNA by Diane
Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
58
REFERENCES
Uses of DNA Extraction | eHowcom httpwwwehowcomabout_5344428_uses-dna-extractionhtmlixzz1MkWQB2TZ
Science 10 A Contextual Approach Heineman Queensland Science Projects Regan Spence Maggie Spenceley
httpenwikipediaorgwikiMolecular_biology httpwwwclinchemorgcgicontent
extract44102201 http
employeescsbsjueduhjakubowskiclassesch331dnaoldnastructurehtml
httpwwwgenomebccaeducationteachersclassroom-activitiesdna-extraction
32
THE WHYrsquoS Salt and Detergents are used to break down
cell walls and nuclear membranes to release the DNA
They work by chemically poking holes in the cell membranes or walls
Once holes are poked in the membranes the membranes can be further disrupted mechanically as with a blender
After that it is easier to get the contents of the cell out including the DNA
LETS HAVE A LOOK AT THE CELL
33
CELL TO DNA
httplearngeneticsutaheducontentlabsextractionhowtoHOW TO EXTRACT DNA FROM ANYTHING LIVING
34
THE WHYrsquoS WHY DETERGENT
Each cell is surrounded by a sack (the cell membrane)
DNA is found inside the second sack (nucleus) within each cell
To see the DNA we have to break it open A cell membrane has 2 layers of lipid(fat)
molecules with protein going through them When the lysis buffer (detergent) comes
close to the cell it captures the lipids and the proteins - breaks open the cell destroying the fatty membrane - DNA now released into the solution
35
A CELLS MEMBRANES HAVE TWO LAYERS OF LIPID (FAT) MOLECULES WITH PROTEINS GOING THROUGH THEM
36
Why detergent How does detergent work
Think about why you use soap to wash dishes or your hands To remove grease and dirt right
Soap molecules and grease molecules are made of two parts
1(Blue) Heads which like water 2(Green) Tails which hate water
37
AFTER ADDING THE DETERGENT WHAT DO YOU HAVE IN YOUR SOUP
38
THE WHYrsquoS ENZYME (MEAT TENDERIZER)
The DNA in the nucleus of the cell is molded folded and protected by proteins The tenderizer cut the proteins away from the DNA
In this experiment meat tenderizer acts as an enzyme to cut proteins just like a pair of scissors
The meat tenderizer cuts the proteins away from the DNA
39
THE WHYrsquoS CUTTING PROTEIN AND DNA
The DNA in the nucleus of the cell is moulded folded and protected by proteins
40
THE WHYrsquoSALCOHOL Alcohol is less dense than water so it floats
on top Look for clumps of white stringy stuff where the water and alcohol layers meet
DNA is not soluble in alcohol - other cell parts are
By adding alcohol DNA precipitates out of the solution and collect at the interface of the alcohol and soap layer
The colder the alcohol the less soluble the DNA will be in it
41
COLD ALCOHOL
DNA dissolves in water but precipitates in alcohol
Cold alcohol is used to separate DNA out of water-based solutions
This allows the DNA to be purified for subsequent genetic testing
Adding alcohol to a solution containing DNA is a simple way to obtain the pure DNA required and colder temperatures slow down enzymes that can break down DNA giving better extraction results
42
WHY IS DNA EXTRACTION IMPORTANT DNA extraction is an important molecular
biology procedure
By definition extraction is taking DNA out of any type of cell for the purpose of analysis
See Handouts for more information on the use of DNA extracted (extension only)
43
SUMMARIZEDNA EXTRACTING FROM KIWI FRUIT
44
BREAK DOWN CELL WALLS
Cells walls have to be destroyed to reach the DNA within cells
Extraction procedures to obtain pure DNA have to get rid of all the molecular and chemical components of the tissue from which the DNA is being extracted
First steps involve lysing or destroying the cell walls This can be done with a variety of chemical agents that are caustic to cell membranes but do not harm the DNA Cells can also be sonicated homogenized or ground up to destroy membranes
45
REMOVING Removing Lipids
Once the cell walls have been destroyed a detergent is added to get rid of the fats and oils that make up the cell membranes Detergents cause the fats and oils to dissolve into the solution
Remove ProteinsProteins and enzymes can be digested by
adding a protease to the solution Proteases break down proteins into small peptides and amino acids
46
PRECIPITATING AND PURIFYING Precipitate DNA
Adding cold alcohol to a solution will cause DNA to precipitate and aggregate The DNA can be collected by centrifuging the sample and pouring off the liquid layer The DNA should exist as a small pellet in the bottom of the centrifuge tube
Purify DNAThe DNA can be washed by re-suspending it
in a cold alcohol solution and re-centrifuging it several times to obtain a very pure DNA sample Typical alcohols used to precipitate DNA include ethanol and Isopropanol This process leaves a very pure sample for stringent DNA testing
47
PART B OBSERVING EXTRACTED DNA UNDER THE MICROSCOPE
48
PART B A CLOSER LOOK
8 Use a dropping pipette to carefully remove some of the threadlike substance from the top of your preparation
9 Place one or two drops onto the middle of a
microscope slide
10 Add two drops of methylene blue Wait 3 or 4 minutes to allow the methylene blue to be absorbed by the DNA
11 Carefully place a cover slip on the slide Gently press
a folded piece of paper towelling over the top of the prepared slide to soak up any excess liquid
12 Observe the DNA under low power then high power
49
DISCUSSIONANSWER IN GROUP RELATED WORKED 1 Write a detailed description of the material
floating at the top of the test tube after the alcohol was added
2 Describe the DNA as it appears under the
microscope under high power 3 Prepare a diagram of your DNA specimen 4 What was the reason for using methylene
blue
50
HOMEWORKCONSTRUCT A FLOW CHART DRAWING MIND MAP OF THE PROCESS YOU USED EXTRACTING THE DNA FROM KIWIrsquoS
Next to each step explain how the method you used was important (Hint What substances make up the membranes of cells and cell organelles
How do detergents work
What is the active ingredient in meat tenderiser
51
TIPS STEPS FOR FLOW CHART In order to release the DNA from the
nuclei of the pea cells you first separated the cells from one another
Then the cell and nuclear membranes needed to be ruptured to release the cell contents and the contents of the nucleus
Once removed from the nuclear membrane the DNA had to be untangled into the visible threadlike structures you ended up with
52
HOMEWORK EXTENSION GROUP QUESTIONS Where can DNA be found in the cell Discuss the action of the soap (detergent) on the cell
What is the purpose of the soap in this activity What was the purpose of the Sodium Chloride Include a
discussion of polarity and charged particles Why was the cold ethanol added to the soap and salt
mixture Describe the appearance of your final product Draw a diagram of DNA containing 5 sets of nucleotide
bases labelling the hydrogen bonds between the bases References and Resources Adapted from Berry Full
of DNA by Diane Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
53
HTTPWWWGENOMEBCCAEDUCATIONTEACHERSCLASSROOM-ACTIVITIESDNA-EXTRACTION
54
EXTRACHROMOSOMES EXIST IN PAIRS Because our chromosomes exist in pairs (and consequently we
have 2 alleles of each gene) we are a diploid species This is why our somatic cells are represented as 2n Our gametes (sperm and ova) on the other hand are haploid and are represented as n Other species may have different ploidy for example
triploid (3n) seedless watermelons tetaploid (4n) salmonidae fish pentaploid (5n) Kenai birch hexaploid (6n) some types of wheat kiwi fruit octaploid (8n) acipenser (a genus
of sturgeon fish strawberies) decaploid (10n) some strawberries dodecaploid (12n) some types of amphibians eg Xenopus
ruwenzoriensis
wwwcyberusca
55
YOUR EXTRACTED DNA UNDER THE MICROSCOPE LOOKED
LIKE
Collaboration-Brainstorming on what to look at under the
- DNA extraction- DNA is too small for even a microscope to see and after the extraction it just appears like a blob True or false
Could you any DNA strand
wwwemployeescsbsjuedu
56
HANDOUT ON DNA EXTRACTIONBACKGROUND EVERY DAY LIFE
EXTRA READING MATERIAL(EXTENSION )
DISCOVERING DNA DNA ON THE INSIDE USE OF DNA EXTRACTION INN EVERYDAY LIFE WHY IS DNA TESTING GOOD MOLECULAR BIOLOGY PCR-based diagnostics of genomic DNA
57
REFERENCES httpwwwsquidoocomhow-to-get-dna-from-a-kiwi-fruit Why Is DNA Testing Good | eHowcom
httpwwwehowcomabout_6684410_dna-testing-good_htmlixzz1MkbcIJs5 Why Is DNA Extraction Important | eHowcom
httpwwwehowcomlist_5839095_dna-extraction-important_htmlixzz1MkZDrqyY
Why Is Sodium Used in DNA Extraction | eHowcom httpwwwehowcomabout_6504902_sodium-used-dna-extraction_htmlixzz1MkaGWg57
Why Is Cold Alcohol Used in DNA Tests | eHowcom httpwwwehowcomabout_6399349_cold-alcohol-used-dna-tests_htmlixzz1MkbEmEdu
httplearngeneticsutaheducontentlabsextractionhowto www kamibudaksainsblogspotcom
httpwwwchemwatch goldcom References and Resources Adapted from Berry Full of DNA by Diane
Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
58
REFERENCES
Uses of DNA Extraction | eHowcom httpwwwehowcomabout_5344428_uses-dna-extractionhtmlixzz1MkWQB2TZ
Science 10 A Contextual Approach Heineman Queensland Science Projects Regan Spence Maggie Spenceley
httpenwikipediaorgwikiMolecular_biology httpwwwclinchemorgcgicontent
extract44102201 http
employeescsbsjueduhjakubowskiclassesch331dnaoldnastructurehtml
httpwwwgenomebccaeducationteachersclassroom-activitiesdna-extraction
33
CELL TO DNA
httplearngeneticsutaheducontentlabsextractionhowtoHOW TO EXTRACT DNA FROM ANYTHING LIVING
34
THE WHYrsquoS WHY DETERGENT
Each cell is surrounded by a sack (the cell membrane)
DNA is found inside the second sack (nucleus) within each cell
To see the DNA we have to break it open A cell membrane has 2 layers of lipid(fat)
molecules with protein going through them When the lysis buffer (detergent) comes
close to the cell it captures the lipids and the proteins - breaks open the cell destroying the fatty membrane - DNA now released into the solution
35
A CELLS MEMBRANES HAVE TWO LAYERS OF LIPID (FAT) MOLECULES WITH PROTEINS GOING THROUGH THEM
36
Why detergent How does detergent work
Think about why you use soap to wash dishes or your hands To remove grease and dirt right
Soap molecules and grease molecules are made of two parts
1(Blue) Heads which like water 2(Green) Tails which hate water
37
AFTER ADDING THE DETERGENT WHAT DO YOU HAVE IN YOUR SOUP
38
THE WHYrsquoS ENZYME (MEAT TENDERIZER)
The DNA in the nucleus of the cell is molded folded and protected by proteins The tenderizer cut the proteins away from the DNA
In this experiment meat tenderizer acts as an enzyme to cut proteins just like a pair of scissors
The meat tenderizer cuts the proteins away from the DNA
39
THE WHYrsquoS CUTTING PROTEIN AND DNA
The DNA in the nucleus of the cell is moulded folded and protected by proteins
40
THE WHYrsquoSALCOHOL Alcohol is less dense than water so it floats
on top Look for clumps of white stringy stuff where the water and alcohol layers meet
DNA is not soluble in alcohol - other cell parts are
By adding alcohol DNA precipitates out of the solution and collect at the interface of the alcohol and soap layer
The colder the alcohol the less soluble the DNA will be in it
41
COLD ALCOHOL
DNA dissolves in water but precipitates in alcohol
Cold alcohol is used to separate DNA out of water-based solutions
This allows the DNA to be purified for subsequent genetic testing
Adding alcohol to a solution containing DNA is a simple way to obtain the pure DNA required and colder temperatures slow down enzymes that can break down DNA giving better extraction results
42
WHY IS DNA EXTRACTION IMPORTANT DNA extraction is an important molecular
biology procedure
By definition extraction is taking DNA out of any type of cell for the purpose of analysis
See Handouts for more information on the use of DNA extracted (extension only)
43
SUMMARIZEDNA EXTRACTING FROM KIWI FRUIT
44
BREAK DOWN CELL WALLS
Cells walls have to be destroyed to reach the DNA within cells
Extraction procedures to obtain pure DNA have to get rid of all the molecular and chemical components of the tissue from which the DNA is being extracted
First steps involve lysing or destroying the cell walls This can be done with a variety of chemical agents that are caustic to cell membranes but do not harm the DNA Cells can also be sonicated homogenized or ground up to destroy membranes
45
REMOVING Removing Lipids
Once the cell walls have been destroyed a detergent is added to get rid of the fats and oils that make up the cell membranes Detergents cause the fats and oils to dissolve into the solution
Remove ProteinsProteins and enzymes can be digested by
adding a protease to the solution Proteases break down proteins into small peptides and amino acids
46
PRECIPITATING AND PURIFYING Precipitate DNA
Adding cold alcohol to a solution will cause DNA to precipitate and aggregate The DNA can be collected by centrifuging the sample and pouring off the liquid layer The DNA should exist as a small pellet in the bottom of the centrifuge tube
Purify DNAThe DNA can be washed by re-suspending it
in a cold alcohol solution and re-centrifuging it several times to obtain a very pure DNA sample Typical alcohols used to precipitate DNA include ethanol and Isopropanol This process leaves a very pure sample for stringent DNA testing
47
PART B OBSERVING EXTRACTED DNA UNDER THE MICROSCOPE
48
PART B A CLOSER LOOK
8 Use a dropping pipette to carefully remove some of the threadlike substance from the top of your preparation
9 Place one or two drops onto the middle of a
microscope slide
10 Add two drops of methylene blue Wait 3 or 4 minutes to allow the methylene blue to be absorbed by the DNA
11 Carefully place a cover slip on the slide Gently press
a folded piece of paper towelling over the top of the prepared slide to soak up any excess liquid
12 Observe the DNA under low power then high power
49
DISCUSSIONANSWER IN GROUP RELATED WORKED 1 Write a detailed description of the material
floating at the top of the test tube after the alcohol was added
2 Describe the DNA as it appears under the
microscope under high power 3 Prepare a diagram of your DNA specimen 4 What was the reason for using methylene
blue
50
HOMEWORKCONSTRUCT A FLOW CHART DRAWING MIND MAP OF THE PROCESS YOU USED EXTRACTING THE DNA FROM KIWIrsquoS
Next to each step explain how the method you used was important (Hint What substances make up the membranes of cells and cell organelles
How do detergents work
What is the active ingredient in meat tenderiser
51
TIPS STEPS FOR FLOW CHART In order to release the DNA from the
nuclei of the pea cells you first separated the cells from one another
Then the cell and nuclear membranes needed to be ruptured to release the cell contents and the contents of the nucleus
Once removed from the nuclear membrane the DNA had to be untangled into the visible threadlike structures you ended up with
52
HOMEWORK EXTENSION GROUP QUESTIONS Where can DNA be found in the cell Discuss the action of the soap (detergent) on the cell
What is the purpose of the soap in this activity What was the purpose of the Sodium Chloride Include a
discussion of polarity and charged particles Why was the cold ethanol added to the soap and salt
mixture Describe the appearance of your final product Draw a diagram of DNA containing 5 sets of nucleotide
bases labelling the hydrogen bonds between the bases References and Resources Adapted from Berry Full
of DNA by Diane Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
53
HTTPWWWGENOMEBCCAEDUCATIONTEACHERSCLASSROOM-ACTIVITIESDNA-EXTRACTION
54
EXTRACHROMOSOMES EXIST IN PAIRS Because our chromosomes exist in pairs (and consequently we
have 2 alleles of each gene) we are a diploid species This is why our somatic cells are represented as 2n Our gametes (sperm and ova) on the other hand are haploid and are represented as n Other species may have different ploidy for example
triploid (3n) seedless watermelons tetaploid (4n) salmonidae fish pentaploid (5n) Kenai birch hexaploid (6n) some types of wheat kiwi fruit octaploid (8n) acipenser (a genus
of sturgeon fish strawberies) decaploid (10n) some strawberries dodecaploid (12n) some types of amphibians eg Xenopus
ruwenzoriensis
wwwcyberusca
55
YOUR EXTRACTED DNA UNDER THE MICROSCOPE LOOKED
LIKE
Collaboration-Brainstorming on what to look at under the
- DNA extraction- DNA is too small for even a microscope to see and after the extraction it just appears like a blob True or false
Could you any DNA strand
wwwemployeescsbsjuedu
56
HANDOUT ON DNA EXTRACTIONBACKGROUND EVERY DAY LIFE
EXTRA READING MATERIAL(EXTENSION )
DISCOVERING DNA DNA ON THE INSIDE USE OF DNA EXTRACTION INN EVERYDAY LIFE WHY IS DNA TESTING GOOD MOLECULAR BIOLOGY PCR-based diagnostics of genomic DNA
57
REFERENCES httpwwwsquidoocomhow-to-get-dna-from-a-kiwi-fruit Why Is DNA Testing Good | eHowcom
httpwwwehowcomabout_6684410_dna-testing-good_htmlixzz1MkbcIJs5 Why Is DNA Extraction Important | eHowcom
httpwwwehowcomlist_5839095_dna-extraction-important_htmlixzz1MkZDrqyY
Why Is Sodium Used in DNA Extraction | eHowcom httpwwwehowcomabout_6504902_sodium-used-dna-extraction_htmlixzz1MkaGWg57
Why Is Cold Alcohol Used in DNA Tests | eHowcom httpwwwehowcomabout_6399349_cold-alcohol-used-dna-tests_htmlixzz1MkbEmEdu
httplearngeneticsutaheducontentlabsextractionhowto www kamibudaksainsblogspotcom
httpwwwchemwatch goldcom References and Resources Adapted from Berry Full of DNA by Diane
Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
58
REFERENCES
Uses of DNA Extraction | eHowcom httpwwwehowcomabout_5344428_uses-dna-extractionhtmlixzz1MkWQB2TZ
Science 10 A Contextual Approach Heineman Queensland Science Projects Regan Spence Maggie Spenceley
httpenwikipediaorgwikiMolecular_biology httpwwwclinchemorgcgicontent
extract44102201 http
employeescsbsjueduhjakubowskiclassesch331dnaoldnastructurehtml
httpwwwgenomebccaeducationteachersclassroom-activitiesdna-extraction
34
THE WHYrsquoS WHY DETERGENT
Each cell is surrounded by a sack (the cell membrane)
DNA is found inside the second sack (nucleus) within each cell
To see the DNA we have to break it open A cell membrane has 2 layers of lipid(fat)
molecules with protein going through them When the lysis buffer (detergent) comes
close to the cell it captures the lipids and the proteins - breaks open the cell destroying the fatty membrane - DNA now released into the solution
35
A CELLS MEMBRANES HAVE TWO LAYERS OF LIPID (FAT) MOLECULES WITH PROTEINS GOING THROUGH THEM
36
Why detergent How does detergent work
Think about why you use soap to wash dishes or your hands To remove grease and dirt right
Soap molecules and grease molecules are made of two parts
1(Blue) Heads which like water 2(Green) Tails which hate water
37
AFTER ADDING THE DETERGENT WHAT DO YOU HAVE IN YOUR SOUP
38
THE WHYrsquoS ENZYME (MEAT TENDERIZER)
The DNA in the nucleus of the cell is molded folded and protected by proteins The tenderizer cut the proteins away from the DNA
In this experiment meat tenderizer acts as an enzyme to cut proteins just like a pair of scissors
The meat tenderizer cuts the proteins away from the DNA
39
THE WHYrsquoS CUTTING PROTEIN AND DNA
The DNA in the nucleus of the cell is moulded folded and protected by proteins
40
THE WHYrsquoSALCOHOL Alcohol is less dense than water so it floats
on top Look for clumps of white stringy stuff where the water and alcohol layers meet
DNA is not soluble in alcohol - other cell parts are
By adding alcohol DNA precipitates out of the solution and collect at the interface of the alcohol and soap layer
The colder the alcohol the less soluble the DNA will be in it
41
COLD ALCOHOL
DNA dissolves in water but precipitates in alcohol
Cold alcohol is used to separate DNA out of water-based solutions
This allows the DNA to be purified for subsequent genetic testing
Adding alcohol to a solution containing DNA is a simple way to obtain the pure DNA required and colder temperatures slow down enzymes that can break down DNA giving better extraction results
42
WHY IS DNA EXTRACTION IMPORTANT DNA extraction is an important molecular
biology procedure
By definition extraction is taking DNA out of any type of cell for the purpose of analysis
See Handouts for more information on the use of DNA extracted (extension only)
43
SUMMARIZEDNA EXTRACTING FROM KIWI FRUIT
44
BREAK DOWN CELL WALLS
Cells walls have to be destroyed to reach the DNA within cells
Extraction procedures to obtain pure DNA have to get rid of all the molecular and chemical components of the tissue from which the DNA is being extracted
First steps involve lysing or destroying the cell walls This can be done with a variety of chemical agents that are caustic to cell membranes but do not harm the DNA Cells can also be sonicated homogenized or ground up to destroy membranes
45
REMOVING Removing Lipids
Once the cell walls have been destroyed a detergent is added to get rid of the fats and oils that make up the cell membranes Detergents cause the fats and oils to dissolve into the solution
Remove ProteinsProteins and enzymes can be digested by
adding a protease to the solution Proteases break down proteins into small peptides and amino acids
46
PRECIPITATING AND PURIFYING Precipitate DNA
Adding cold alcohol to a solution will cause DNA to precipitate and aggregate The DNA can be collected by centrifuging the sample and pouring off the liquid layer The DNA should exist as a small pellet in the bottom of the centrifuge tube
Purify DNAThe DNA can be washed by re-suspending it
in a cold alcohol solution and re-centrifuging it several times to obtain a very pure DNA sample Typical alcohols used to precipitate DNA include ethanol and Isopropanol This process leaves a very pure sample for stringent DNA testing
47
PART B OBSERVING EXTRACTED DNA UNDER THE MICROSCOPE
48
PART B A CLOSER LOOK
8 Use a dropping pipette to carefully remove some of the threadlike substance from the top of your preparation
9 Place one or two drops onto the middle of a
microscope slide
10 Add two drops of methylene blue Wait 3 or 4 minutes to allow the methylene blue to be absorbed by the DNA
11 Carefully place a cover slip on the slide Gently press
a folded piece of paper towelling over the top of the prepared slide to soak up any excess liquid
12 Observe the DNA under low power then high power
49
DISCUSSIONANSWER IN GROUP RELATED WORKED 1 Write a detailed description of the material
floating at the top of the test tube after the alcohol was added
2 Describe the DNA as it appears under the
microscope under high power 3 Prepare a diagram of your DNA specimen 4 What was the reason for using methylene
blue
50
HOMEWORKCONSTRUCT A FLOW CHART DRAWING MIND MAP OF THE PROCESS YOU USED EXTRACTING THE DNA FROM KIWIrsquoS
Next to each step explain how the method you used was important (Hint What substances make up the membranes of cells and cell organelles
How do detergents work
What is the active ingredient in meat tenderiser
51
TIPS STEPS FOR FLOW CHART In order to release the DNA from the
nuclei of the pea cells you first separated the cells from one another
Then the cell and nuclear membranes needed to be ruptured to release the cell contents and the contents of the nucleus
Once removed from the nuclear membrane the DNA had to be untangled into the visible threadlike structures you ended up with
52
HOMEWORK EXTENSION GROUP QUESTIONS Where can DNA be found in the cell Discuss the action of the soap (detergent) on the cell
What is the purpose of the soap in this activity What was the purpose of the Sodium Chloride Include a
discussion of polarity and charged particles Why was the cold ethanol added to the soap and salt
mixture Describe the appearance of your final product Draw a diagram of DNA containing 5 sets of nucleotide
bases labelling the hydrogen bonds between the bases References and Resources Adapted from Berry Full
of DNA by Diane Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
53
HTTPWWWGENOMEBCCAEDUCATIONTEACHERSCLASSROOM-ACTIVITIESDNA-EXTRACTION
54
EXTRACHROMOSOMES EXIST IN PAIRS Because our chromosomes exist in pairs (and consequently we
have 2 alleles of each gene) we are a diploid species This is why our somatic cells are represented as 2n Our gametes (sperm and ova) on the other hand are haploid and are represented as n Other species may have different ploidy for example
triploid (3n) seedless watermelons tetaploid (4n) salmonidae fish pentaploid (5n) Kenai birch hexaploid (6n) some types of wheat kiwi fruit octaploid (8n) acipenser (a genus
of sturgeon fish strawberies) decaploid (10n) some strawberries dodecaploid (12n) some types of amphibians eg Xenopus
ruwenzoriensis
wwwcyberusca
55
YOUR EXTRACTED DNA UNDER THE MICROSCOPE LOOKED
LIKE
Collaboration-Brainstorming on what to look at under the
- DNA extraction- DNA is too small for even a microscope to see and after the extraction it just appears like a blob True or false
Could you any DNA strand
wwwemployeescsbsjuedu
56
HANDOUT ON DNA EXTRACTIONBACKGROUND EVERY DAY LIFE
EXTRA READING MATERIAL(EXTENSION )
DISCOVERING DNA DNA ON THE INSIDE USE OF DNA EXTRACTION INN EVERYDAY LIFE WHY IS DNA TESTING GOOD MOLECULAR BIOLOGY PCR-based diagnostics of genomic DNA
57
REFERENCES httpwwwsquidoocomhow-to-get-dna-from-a-kiwi-fruit Why Is DNA Testing Good | eHowcom
httpwwwehowcomabout_6684410_dna-testing-good_htmlixzz1MkbcIJs5 Why Is DNA Extraction Important | eHowcom
httpwwwehowcomlist_5839095_dna-extraction-important_htmlixzz1MkZDrqyY
Why Is Sodium Used in DNA Extraction | eHowcom httpwwwehowcomabout_6504902_sodium-used-dna-extraction_htmlixzz1MkaGWg57
Why Is Cold Alcohol Used in DNA Tests | eHowcom httpwwwehowcomabout_6399349_cold-alcohol-used-dna-tests_htmlixzz1MkbEmEdu
httplearngeneticsutaheducontentlabsextractionhowto www kamibudaksainsblogspotcom
httpwwwchemwatch goldcom References and Resources Adapted from Berry Full of DNA by Diane
Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
58
REFERENCES
Uses of DNA Extraction | eHowcom httpwwwehowcomabout_5344428_uses-dna-extractionhtmlixzz1MkWQB2TZ
Science 10 A Contextual Approach Heineman Queensland Science Projects Regan Spence Maggie Spenceley
httpenwikipediaorgwikiMolecular_biology httpwwwclinchemorgcgicontent
extract44102201 http
employeescsbsjueduhjakubowskiclassesch331dnaoldnastructurehtml
httpwwwgenomebccaeducationteachersclassroom-activitiesdna-extraction
35
A CELLS MEMBRANES HAVE TWO LAYERS OF LIPID (FAT) MOLECULES WITH PROTEINS GOING THROUGH THEM
36
Why detergent How does detergent work
Think about why you use soap to wash dishes or your hands To remove grease and dirt right
Soap molecules and grease molecules are made of two parts
1(Blue) Heads which like water 2(Green) Tails which hate water
37
AFTER ADDING THE DETERGENT WHAT DO YOU HAVE IN YOUR SOUP
38
THE WHYrsquoS ENZYME (MEAT TENDERIZER)
The DNA in the nucleus of the cell is molded folded and protected by proteins The tenderizer cut the proteins away from the DNA
In this experiment meat tenderizer acts as an enzyme to cut proteins just like a pair of scissors
The meat tenderizer cuts the proteins away from the DNA
39
THE WHYrsquoS CUTTING PROTEIN AND DNA
The DNA in the nucleus of the cell is moulded folded and protected by proteins
40
THE WHYrsquoSALCOHOL Alcohol is less dense than water so it floats
on top Look for clumps of white stringy stuff where the water and alcohol layers meet
DNA is not soluble in alcohol - other cell parts are
By adding alcohol DNA precipitates out of the solution and collect at the interface of the alcohol and soap layer
The colder the alcohol the less soluble the DNA will be in it
41
COLD ALCOHOL
DNA dissolves in water but precipitates in alcohol
Cold alcohol is used to separate DNA out of water-based solutions
This allows the DNA to be purified for subsequent genetic testing
Adding alcohol to a solution containing DNA is a simple way to obtain the pure DNA required and colder temperatures slow down enzymes that can break down DNA giving better extraction results
42
WHY IS DNA EXTRACTION IMPORTANT DNA extraction is an important molecular
biology procedure
By definition extraction is taking DNA out of any type of cell for the purpose of analysis
See Handouts for more information on the use of DNA extracted (extension only)
43
SUMMARIZEDNA EXTRACTING FROM KIWI FRUIT
44
BREAK DOWN CELL WALLS
Cells walls have to be destroyed to reach the DNA within cells
Extraction procedures to obtain pure DNA have to get rid of all the molecular and chemical components of the tissue from which the DNA is being extracted
First steps involve lysing or destroying the cell walls This can be done with a variety of chemical agents that are caustic to cell membranes but do not harm the DNA Cells can also be sonicated homogenized or ground up to destroy membranes
45
REMOVING Removing Lipids
Once the cell walls have been destroyed a detergent is added to get rid of the fats and oils that make up the cell membranes Detergents cause the fats and oils to dissolve into the solution
Remove ProteinsProteins and enzymes can be digested by
adding a protease to the solution Proteases break down proteins into small peptides and amino acids
46
PRECIPITATING AND PURIFYING Precipitate DNA
Adding cold alcohol to a solution will cause DNA to precipitate and aggregate The DNA can be collected by centrifuging the sample and pouring off the liquid layer The DNA should exist as a small pellet in the bottom of the centrifuge tube
Purify DNAThe DNA can be washed by re-suspending it
in a cold alcohol solution and re-centrifuging it several times to obtain a very pure DNA sample Typical alcohols used to precipitate DNA include ethanol and Isopropanol This process leaves a very pure sample for stringent DNA testing
47
PART B OBSERVING EXTRACTED DNA UNDER THE MICROSCOPE
48
PART B A CLOSER LOOK
8 Use a dropping pipette to carefully remove some of the threadlike substance from the top of your preparation
9 Place one or two drops onto the middle of a
microscope slide
10 Add two drops of methylene blue Wait 3 or 4 minutes to allow the methylene blue to be absorbed by the DNA
11 Carefully place a cover slip on the slide Gently press
a folded piece of paper towelling over the top of the prepared slide to soak up any excess liquid
12 Observe the DNA under low power then high power
49
DISCUSSIONANSWER IN GROUP RELATED WORKED 1 Write a detailed description of the material
floating at the top of the test tube after the alcohol was added
2 Describe the DNA as it appears under the
microscope under high power 3 Prepare a diagram of your DNA specimen 4 What was the reason for using methylene
blue
50
HOMEWORKCONSTRUCT A FLOW CHART DRAWING MIND MAP OF THE PROCESS YOU USED EXTRACTING THE DNA FROM KIWIrsquoS
Next to each step explain how the method you used was important (Hint What substances make up the membranes of cells and cell organelles
How do detergents work
What is the active ingredient in meat tenderiser
51
TIPS STEPS FOR FLOW CHART In order to release the DNA from the
nuclei of the pea cells you first separated the cells from one another
Then the cell and nuclear membranes needed to be ruptured to release the cell contents and the contents of the nucleus
Once removed from the nuclear membrane the DNA had to be untangled into the visible threadlike structures you ended up with
52
HOMEWORK EXTENSION GROUP QUESTIONS Where can DNA be found in the cell Discuss the action of the soap (detergent) on the cell
What is the purpose of the soap in this activity What was the purpose of the Sodium Chloride Include a
discussion of polarity and charged particles Why was the cold ethanol added to the soap and salt
mixture Describe the appearance of your final product Draw a diagram of DNA containing 5 sets of nucleotide
bases labelling the hydrogen bonds between the bases References and Resources Adapted from Berry Full
of DNA by Diane Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
53
HTTPWWWGENOMEBCCAEDUCATIONTEACHERSCLASSROOM-ACTIVITIESDNA-EXTRACTION
54
EXTRACHROMOSOMES EXIST IN PAIRS Because our chromosomes exist in pairs (and consequently we
have 2 alleles of each gene) we are a diploid species This is why our somatic cells are represented as 2n Our gametes (sperm and ova) on the other hand are haploid and are represented as n Other species may have different ploidy for example
triploid (3n) seedless watermelons tetaploid (4n) salmonidae fish pentaploid (5n) Kenai birch hexaploid (6n) some types of wheat kiwi fruit octaploid (8n) acipenser (a genus
of sturgeon fish strawberies) decaploid (10n) some strawberries dodecaploid (12n) some types of amphibians eg Xenopus
ruwenzoriensis
wwwcyberusca
55
YOUR EXTRACTED DNA UNDER THE MICROSCOPE LOOKED
LIKE
Collaboration-Brainstorming on what to look at under the
- DNA extraction- DNA is too small for even a microscope to see and after the extraction it just appears like a blob True or false
Could you any DNA strand
wwwemployeescsbsjuedu
56
HANDOUT ON DNA EXTRACTIONBACKGROUND EVERY DAY LIFE
EXTRA READING MATERIAL(EXTENSION )
DISCOVERING DNA DNA ON THE INSIDE USE OF DNA EXTRACTION INN EVERYDAY LIFE WHY IS DNA TESTING GOOD MOLECULAR BIOLOGY PCR-based diagnostics of genomic DNA
57
REFERENCES httpwwwsquidoocomhow-to-get-dna-from-a-kiwi-fruit Why Is DNA Testing Good | eHowcom
httpwwwehowcomabout_6684410_dna-testing-good_htmlixzz1MkbcIJs5 Why Is DNA Extraction Important | eHowcom
httpwwwehowcomlist_5839095_dna-extraction-important_htmlixzz1MkZDrqyY
Why Is Sodium Used in DNA Extraction | eHowcom httpwwwehowcomabout_6504902_sodium-used-dna-extraction_htmlixzz1MkaGWg57
Why Is Cold Alcohol Used in DNA Tests | eHowcom httpwwwehowcomabout_6399349_cold-alcohol-used-dna-tests_htmlixzz1MkbEmEdu
httplearngeneticsutaheducontentlabsextractionhowto www kamibudaksainsblogspotcom
httpwwwchemwatch goldcom References and Resources Adapted from Berry Full of DNA by Diane
Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
58
REFERENCES
Uses of DNA Extraction | eHowcom httpwwwehowcomabout_5344428_uses-dna-extractionhtmlixzz1MkWQB2TZ
Science 10 A Contextual Approach Heineman Queensland Science Projects Regan Spence Maggie Spenceley
httpenwikipediaorgwikiMolecular_biology httpwwwclinchemorgcgicontent
extract44102201 http
employeescsbsjueduhjakubowskiclassesch331dnaoldnastructurehtml
httpwwwgenomebccaeducationteachersclassroom-activitiesdna-extraction
36
Why detergent How does detergent work
Think about why you use soap to wash dishes or your hands To remove grease and dirt right
Soap molecules and grease molecules are made of two parts
1(Blue) Heads which like water 2(Green) Tails which hate water
37
AFTER ADDING THE DETERGENT WHAT DO YOU HAVE IN YOUR SOUP
38
THE WHYrsquoS ENZYME (MEAT TENDERIZER)
The DNA in the nucleus of the cell is molded folded and protected by proteins The tenderizer cut the proteins away from the DNA
In this experiment meat tenderizer acts as an enzyme to cut proteins just like a pair of scissors
The meat tenderizer cuts the proteins away from the DNA
39
THE WHYrsquoS CUTTING PROTEIN AND DNA
The DNA in the nucleus of the cell is moulded folded and protected by proteins
40
THE WHYrsquoSALCOHOL Alcohol is less dense than water so it floats
on top Look for clumps of white stringy stuff where the water and alcohol layers meet
DNA is not soluble in alcohol - other cell parts are
By adding alcohol DNA precipitates out of the solution and collect at the interface of the alcohol and soap layer
The colder the alcohol the less soluble the DNA will be in it
41
COLD ALCOHOL
DNA dissolves in water but precipitates in alcohol
Cold alcohol is used to separate DNA out of water-based solutions
This allows the DNA to be purified for subsequent genetic testing
Adding alcohol to a solution containing DNA is a simple way to obtain the pure DNA required and colder temperatures slow down enzymes that can break down DNA giving better extraction results
42
WHY IS DNA EXTRACTION IMPORTANT DNA extraction is an important molecular
biology procedure
By definition extraction is taking DNA out of any type of cell for the purpose of analysis
See Handouts for more information on the use of DNA extracted (extension only)
43
SUMMARIZEDNA EXTRACTING FROM KIWI FRUIT
44
BREAK DOWN CELL WALLS
Cells walls have to be destroyed to reach the DNA within cells
Extraction procedures to obtain pure DNA have to get rid of all the molecular and chemical components of the tissue from which the DNA is being extracted
First steps involve lysing or destroying the cell walls This can be done with a variety of chemical agents that are caustic to cell membranes but do not harm the DNA Cells can also be sonicated homogenized or ground up to destroy membranes
45
REMOVING Removing Lipids
Once the cell walls have been destroyed a detergent is added to get rid of the fats and oils that make up the cell membranes Detergents cause the fats and oils to dissolve into the solution
Remove ProteinsProteins and enzymes can be digested by
adding a protease to the solution Proteases break down proteins into small peptides and amino acids
46
PRECIPITATING AND PURIFYING Precipitate DNA
Adding cold alcohol to a solution will cause DNA to precipitate and aggregate The DNA can be collected by centrifuging the sample and pouring off the liquid layer The DNA should exist as a small pellet in the bottom of the centrifuge tube
Purify DNAThe DNA can be washed by re-suspending it
in a cold alcohol solution and re-centrifuging it several times to obtain a very pure DNA sample Typical alcohols used to precipitate DNA include ethanol and Isopropanol This process leaves a very pure sample for stringent DNA testing
47
PART B OBSERVING EXTRACTED DNA UNDER THE MICROSCOPE
48
PART B A CLOSER LOOK
8 Use a dropping pipette to carefully remove some of the threadlike substance from the top of your preparation
9 Place one or two drops onto the middle of a
microscope slide
10 Add two drops of methylene blue Wait 3 or 4 minutes to allow the methylene blue to be absorbed by the DNA
11 Carefully place a cover slip on the slide Gently press
a folded piece of paper towelling over the top of the prepared slide to soak up any excess liquid
12 Observe the DNA under low power then high power
49
DISCUSSIONANSWER IN GROUP RELATED WORKED 1 Write a detailed description of the material
floating at the top of the test tube after the alcohol was added
2 Describe the DNA as it appears under the
microscope under high power 3 Prepare a diagram of your DNA specimen 4 What was the reason for using methylene
blue
50
HOMEWORKCONSTRUCT A FLOW CHART DRAWING MIND MAP OF THE PROCESS YOU USED EXTRACTING THE DNA FROM KIWIrsquoS
Next to each step explain how the method you used was important (Hint What substances make up the membranes of cells and cell organelles
How do detergents work
What is the active ingredient in meat tenderiser
51
TIPS STEPS FOR FLOW CHART In order to release the DNA from the
nuclei of the pea cells you first separated the cells from one another
Then the cell and nuclear membranes needed to be ruptured to release the cell contents and the contents of the nucleus
Once removed from the nuclear membrane the DNA had to be untangled into the visible threadlike structures you ended up with
52
HOMEWORK EXTENSION GROUP QUESTIONS Where can DNA be found in the cell Discuss the action of the soap (detergent) on the cell
What is the purpose of the soap in this activity What was the purpose of the Sodium Chloride Include a
discussion of polarity and charged particles Why was the cold ethanol added to the soap and salt
mixture Describe the appearance of your final product Draw a diagram of DNA containing 5 sets of nucleotide
bases labelling the hydrogen bonds between the bases References and Resources Adapted from Berry Full
of DNA by Diane Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
53
HTTPWWWGENOMEBCCAEDUCATIONTEACHERSCLASSROOM-ACTIVITIESDNA-EXTRACTION
54
EXTRACHROMOSOMES EXIST IN PAIRS Because our chromosomes exist in pairs (and consequently we
have 2 alleles of each gene) we are a diploid species This is why our somatic cells are represented as 2n Our gametes (sperm and ova) on the other hand are haploid and are represented as n Other species may have different ploidy for example
triploid (3n) seedless watermelons tetaploid (4n) salmonidae fish pentaploid (5n) Kenai birch hexaploid (6n) some types of wheat kiwi fruit octaploid (8n) acipenser (a genus
of sturgeon fish strawberies) decaploid (10n) some strawberries dodecaploid (12n) some types of amphibians eg Xenopus
ruwenzoriensis
wwwcyberusca
55
YOUR EXTRACTED DNA UNDER THE MICROSCOPE LOOKED
LIKE
Collaboration-Brainstorming on what to look at under the
- DNA extraction- DNA is too small for even a microscope to see and after the extraction it just appears like a blob True or false
Could you any DNA strand
wwwemployeescsbsjuedu
56
HANDOUT ON DNA EXTRACTIONBACKGROUND EVERY DAY LIFE
EXTRA READING MATERIAL(EXTENSION )
DISCOVERING DNA DNA ON THE INSIDE USE OF DNA EXTRACTION INN EVERYDAY LIFE WHY IS DNA TESTING GOOD MOLECULAR BIOLOGY PCR-based diagnostics of genomic DNA
57
REFERENCES httpwwwsquidoocomhow-to-get-dna-from-a-kiwi-fruit Why Is DNA Testing Good | eHowcom
httpwwwehowcomabout_6684410_dna-testing-good_htmlixzz1MkbcIJs5 Why Is DNA Extraction Important | eHowcom
httpwwwehowcomlist_5839095_dna-extraction-important_htmlixzz1MkZDrqyY
Why Is Sodium Used in DNA Extraction | eHowcom httpwwwehowcomabout_6504902_sodium-used-dna-extraction_htmlixzz1MkaGWg57
Why Is Cold Alcohol Used in DNA Tests | eHowcom httpwwwehowcomabout_6399349_cold-alcohol-used-dna-tests_htmlixzz1MkbEmEdu
httplearngeneticsutaheducontentlabsextractionhowto www kamibudaksainsblogspotcom
httpwwwchemwatch goldcom References and Resources Adapted from Berry Full of DNA by Diane
Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
58
REFERENCES
Uses of DNA Extraction | eHowcom httpwwwehowcomabout_5344428_uses-dna-extractionhtmlixzz1MkWQB2TZ
Science 10 A Contextual Approach Heineman Queensland Science Projects Regan Spence Maggie Spenceley
httpenwikipediaorgwikiMolecular_biology httpwwwclinchemorgcgicontent
extract44102201 http
employeescsbsjueduhjakubowskiclassesch331dnaoldnastructurehtml
httpwwwgenomebccaeducationteachersclassroom-activitiesdna-extraction
37
AFTER ADDING THE DETERGENT WHAT DO YOU HAVE IN YOUR SOUP
38
THE WHYrsquoS ENZYME (MEAT TENDERIZER)
The DNA in the nucleus of the cell is molded folded and protected by proteins The tenderizer cut the proteins away from the DNA
In this experiment meat tenderizer acts as an enzyme to cut proteins just like a pair of scissors
The meat tenderizer cuts the proteins away from the DNA
39
THE WHYrsquoS CUTTING PROTEIN AND DNA
The DNA in the nucleus of the cell is moulded folded and protected by proteins
40
THE WHYrsquoSALCOHOL Alcohol is less dense than water so it floats
on top Look for clumps of white stringy stuff where the water and alcohol layers meet
DNA is not soluble in alcohol - other cell parts are
By adding alcohol DNA precipitates out of the solution and collect at the interface of the alcohol and soap layer
The colder the alcohol the less soluble the DNA will be in it
41
COLD ALCOHOL
DNA dissolves in water but precipitates in alcohol
Cold alcohol is used to separate DNA out of water-based solutions
This allows the DNA to be purified for subsequent genetic testing
Adding alcohol to a solution containing DNA is a simple way to obtain the pure DNA required and colder temperatures slow down enzymes that can break down DNA giving better extraction results
42
WHY IS DNA EXTRACTION IMPORTANT DNA extraction is an important molecular
biology procedure
By definition extraction is taking DNA out of any type of cell for the purpose of analysis
See Handouts for more information on the use of DNA extracted (extension only)
43
SUMMARIZEDNA EXTRACTING FROM KIWI FRUIT
44
BREAK DOWN CELL WALLS
Cells walls have to be destroyed to reach the DNA within cells
Extraction procedures to obtain pure DNA have to get rid of all the molecular and chemical components of the tissue from which the DNA is being extracted
First steps involve lysing or destroying the cell walls This can be done with a variety of chemical agents that are caustic to cell membranes but do not harm the DNA Cells can also be sonicated homogenized or ground up to destroy membranes
45
REMOVING Removing Lipids
Once the cell walls have been destroyed a detergent is added to get rid of the fats and oils that make up the cell membranes Detergents cause the fats and oils to dissolve into the solution
Remove ProteinsProteins and enzymes can be digested by
adding a protease to the solution Proteases break down proteins into small peptides and amino acids
46
PRECIPITATING AND PURIFYING Precipitate DNA
Adding cold alcohol to a solution will cause DNA to precipitate and aggregate The DNA can be collected by centrifuging the sample and pouring off the liquid layer The DNA should exist as a small pellet in the bottom of the centrifuge tube
Purify DNAThe DNA can be washed by re-suspending it
in a cold alcohol solution and re-centrifuging it several times to obtain a very pure DNA sample Typical alcohols used to precipitate DNA include ethanol and Isopropanol This process leaves a very pure sample for stringent DNA testing
47
PART B OBSERVING EXTRACTED DNA UNDER THE MICROSCOPE
48
PART B A CLOSER LOOK
8 Use a dropping pipette to carefully remove some of the threadlike substance from the top of your preparation
9 Place one or two drops onto the middle of a
microscope slide
10 Add two drops of methylene blue Wait 3 or 4 minutes to allow the methylene blue to be absorbed by the DNA
11 Carefully place a cover slip on the slide Gently press
a folded piece of paper towelling over the top of the prepared slide to soak up any excess liquid
12 Observe the DNA under low power then high power
49
DISCUSSIONANSWER IN GROUP RELATED WORKED 1 Write a detailed description of the material
floating at the top of the test tube after the alcohol was added
2 Describe the DNA as it appears under the
microscope under high power 3 Prepare a diagram of your DNA specimen 4 What was the reason for using methylene
blue
50
HOMEWORKCONSTRUCT A FLOW CHART DRAWING MIND MAP OF THE PROCESS YOU USED EXTRACTING THE DNA FROM KIWIrsquoS
Next to each step explain how the method you used was important (Hint What substances make up the membranes of cells and cell organelles
How do detergents work
What is the active ingredient in meat tenderiser
51
TIPS STEPS FOR FLOW CHART In order to release the DNA from the
nuclei of the pea cells you first separated the cells from one another
Then the cell and nuclear membranes needed to be ruptured to release the cell contents and the contents of the nucleus
Once removed from the nuclear membrane the DNA had to be untangled into the visible threadlike structures you ended up with
52
HOMEWORK EXTENSION GROUP QUESTIONS Where can DNA be found in the cell Discuss the action of the soap (detergent) on the cell
What is the purpose of the soap in this activity What was the purpose of the Sodium Chloride Include a
discussion of polarity and charged particles Why was the cold ethanol added to the soap and salt
mixture Describe the appearance of your final product Draw a diagram of DNA containing 5 sets of nucleotide
bases labelling the hydrogen bonds between the bases References and Resources Adapted from Berry Full
of DNA by Diane Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
53
HTTPWWWGENOMEBCCAEDUCATIONTEACHERSCLASSROOM-ACTIVITIESDNA-EXTRACTION
54
EXTRACHROMOSOMES EXIST IN PAIRS Because our chromosomes exist in pairs (and consequently we
have 2 alleles of each gene) we are a diploid species This is why our somatic cells are represented as 2n Our gametes (sperm and ova) on the other hand are haploid and are represented as n Other species may have different ploidy for example
triploid (3n) seedless watermelons tetaploid (4n) salmonidae fish pentaploid (5n) Kenai birch hexaploid (6n) some types of wheat kiwi fruit octaploid (8n) acipenser (a genus
of sturgeon fish strawberies) decaploid (10n) some strawberries dodecaploid (12n) some types of amphibians eg Xenopus
ruwenzoriensis
wwwcyberusca
55
YOUR EXTRACTED DNA UNDER THE MICROSCOPE LOOKED
LIKE
Collaboration-Brainstorming on what to look at under the
- DNA extraction- DNA is too small for even a microscope to see and after the extraction it just appears like a blob True or false
Could you any DNA strand
wwwemployeescsbsjuedu
56
HANDOUT ON DNA EXTRACTIONBACKGROUND EVERY DAY LIFE
EXTRA READING MATERIAL(EXTENSION )
DISCOVERING DNA DNA ON THE INSIDE USE OF DNA EXTRACTION INN EVERYDAY LIFE WHY IS DNA TESTING GOOD MOLECULAR BIOLOGY PCR-based diagnostics of genomic DNA
57
REFERENCES httpwwwsquidoocomhow-to-get-dna-from-a-kiwi-fruit Why Is DNA Testing Good | eHowcom
httpwwwehowcomabout_6684410_dna-testing-good_htmlixzz1MkbcIJs5 Why Is DNA Extraction Important | eHowcom
httpwwwehowcomlist_5839095_dna-extraction-important_htmlixzz1MkZDrqyY
Why Is Sodium Used in DNA Extraction | eHowcom httpwwwehowcomabout_6504902_sodium-used-dna-extraction_htmlixzz1MkaGWg57
Why Is Cold Alcohol Used in DNA Tests | eHowcom httpwwwehowcomabout_6399349_cold-alcohol-used-dna-tests_htmlixzz1MkbEmEdu
httplearngeneticsutaheducontentlabsextractionhowto www kamibudaksainsblogspotcom
httpwwwchemwatch goldcom References and Resources Adapted from Berry Full of DNA by Diane
Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
58
REFERENCES
Uses of DNA Extraction | eHowcom httpwwwehowcomabout_5344428_uses-dna-extractionhtmlixzz1MkWQB2TZ
Science 10 A Contextual Approach Heineman Queensland Science Projects Regan Spence Maggie Spenceley
httpenwikipediaorgwikiMolecular_biology httpwwwclinchemorgcgicontent
extract44102201 http
employeescsbsjueduhjakubowskiclassesch331dnaoldnastructurehtml
httpwwwgenomebccaeducationteachersclassroom-activitiesdna-extraction
38
THE WHYrsquoS ENZYME (MEAT TENDERIZER)
The DNA in the nucleus of the cell is molded folded and protected by proteins The tenderizer cut the proteins away from the DNA
In this experiment meat tenderizer acts as an enzyme to cut proteins just like a pair of scissors
The meat tenderizer cuts the proteins away from the DNA
39
THE WHYrsquoS CUTTING PROTEIN AND DNA
The DNA in the nucleus of the cell is moulded folded and protected by proteins
40
THE WHYrsquoSALCOHOL Alcohol is less dense than water so it floats
on top Look for clumps of white stringy stuff where the water and alcohol layers meet
DNA is not soluble in alcohol - other cell parts are
By adding alcohol DNA precipitates out of the solution and collect at the interface of the alcohol and soap layer
The colder the alcohol the less soluble the DNA will be in it
41
COLD ALCOHOL
DNA dissolves in water but precipitates in alcohol
Cold alcohol is used to separate DNA out of water-based solutions
This allows the DNA to be purified for subsequent genetic testing
Adding alcohol to a solution containing DNA is a simple way to obtain the pure DNA required and colder temperatures slow down enzymes that can break down DNA giving better extraction results
42
WHY IS DNA EXTRACTION IMPORTANT DNA extraction is an important molecular
biology procedure
By definition extraction is taking DNA out of any type of cell for the purpose of analysis
See Handouts for more information on the use of DNA extracted (extension only)
43
SUMMARIZEDNA EXTRACTING FROM KIWI FRUIT
44
BREAK DOWN CELL WALLS
Cells walls have to be destroyed to reach the DNA within cells
Extraction procedures to obtain pure DNA have to get rid of all the molecular and chemical components of the tissue from which the DNA is being extracted
First steps involve lysing or destroying the cell walls This can be done with a variety of chemical agents that are caustic to cell membranes but do not harm the DNA Cells can also be sonicated homogenized or ground up to destroy membranes
45
REMOVING Removing Lipids
Once the cell walls have been destroyed a detergent is added to get rid of the fats and oils that make up the cell membranes Detergents cause the fats and oils to dissolve into the solution
Remove ProteinsProteins and enzymes can be digested by
adding a protease to the solution Proteases break down proteins into small peptides and amino acids
46
PRECIPITATING AND PURIFYING Precipitate DNA
Adding cold alcohol to a solution will cause DNA to precipitate and aggregate The DNA can be collected by centrifuging the sample and pouring off the liquid layer The DNA should exist as a small pellet in the bottom of the centrifuge tube
Purify DNAThe DNA can be washed by re-suspending it
in a cold alcohol solution and re-centrifuging it several times to obtain a very pure DNA sample Typical alcohols used to precipitate DNA include ethanol and Isopropanol This process leaves a very pure sample for stringent DNA testing
47
PART B OBSERVING EXTRACTED DNA UNDER THE MICROSCOPE
48
PART B A CLOSER LOOK
8 Use a dropping pipette to carefully remove some of the threadlike substance from the top of your preparation
9 Place one or two drops onto the middle of a
microscope slide
10 Add two drops of methylene blue Wait 3 or 4 minutes to allow the methylene blue to be absorbed by the DNA
11 Carefully place a cover slip on the slide Gently press
a folded piece of paper towelling over the top of the prepared slide to soak up any excess liquid
12 Observe the DNA under low power then high power
49
DISCUSSIONANSWER IN GROUP RELATED WORKED 1 Write a detailed description of the material
floating at the top of the test tube after the alcohol was added
2 Describe the DNA as it appears under the
microscope under high power 3 Prepare a diagram of your DNA specimen 4 What was the reason for using methylene
blue
50
HOMEWORKCONSTRUCT A FLOW CHART DRAWING MIND MAP OF THE PROCESS YOU USED EXTRACTING THE DNA FROM KIWIrsquoS
Next to each step explain how the method you used was important (Hint What substances make up the membranes of cells and cell organelles
How do detergents work
What is the active ingredient in meat tenderiser
51
TIPS STEPS FOR FLOW CHART In order to release the DNA from the
nuclei of the pea cells you first separated the cells from one another
Then the cell and nuclear membranes needed to be ruptured to release the cell contents and the contents of the nucleus
Once removed from the nuclear membrane the DNA had to be untangled into the visible threadlike structures you ended up with
52
HOMEWORK EXTENSION GROUP QUESTIONS Where can DNA be found in the cell Discuss the action of the soap (detergent) on the cell
What is the purpose of the soap in this activity What was the purpose of the Sodium Chloride Include a
discussion of polarity and charged particles Why was the cold ethanol added to the soap and salt
mixture Describe the appearance of your final product Draw a diagram of DNA containing 5 sets of nucleotide
bases labelling the hydrogen bonds between the bases References and Resources Adapted from Berry Full
of DNA by Diane Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
53
HTTPWWWGENOMEBCCAEDUCATIONTEACHERSCLASSROOM-ACTIVITIESDNA-EXTRACTION
54
EXTRACHROMOSOMES EXIST IN PAIRS Because our chromosomes exist in pairs (and consequently we
have 2 alleles of each gene) we are a diploid species This is why our somatic cells are represented as 2n Our gametes (sperm and ova) on the other hand are haploid and are represented as n Other species may have different ploidy for example
triploid (3n) seedless watermelons tetaploid (4n) salmonidae fish pentaploid (5n) Kenai birch hexaploid (6n) some types of wheat kiwi fruit octaploid (8n) acipenser (a genus
of sturgeon fish strawberies) decaploid (10n) some strawberries dodecaploid (12n) some types of amphibians eg Xenopus
ruwenzoriensis
wwwcyberusca
55
YOUR EXTRACTED DNA UNDER THE MICROSCOPE LOOKED
LIKE
Collaboration-Brainstorming on what to look at under the
- DNA extraction- DNA is too small for even a microscope to see and after the extraction it just appears like a blob True or false
Could you any DNA strand
wwwemployeescsbsjuedu
56
HANDOUT ON DNA EXTRACTIONBACKGROUND EVERY DAY LIFE
EXTRA READING MATERIAL(EXTENSION )
DISCOVERING DNA DNA ON THE INSIDE USE OF DNA EXTRACTION INN EVERYDAY LIFE WHY IS DNA TESTING GOOD MOLECULAR BIOLOGY PCR-based diagnostics of genomic DNA
57
REFERENCES httpwwwsquidoocomhow-to-get-dna-from-a-kiwi-fruit Why Is DNA Testing Good | eHowcom
httpwwwehowcomabout_6684410_dna-testing-good_htmlixzz1MkbcIJs5 Why Is DNA Extraction Important | eHowcom
httpwwwehowcomlist_5839095_dna-extraction-important_htmlixzz1MkZDrqyY
Why Is Sodium Used in DNA Extraction | eHowcom httpwwwehowcomabout_6504902_sodium-used-dna-extraction_htmlixzz1MkaGWg57
Why Is Cold Alcohol Used in DNA Tests | eHowcom httpwwwehowcomabout_6399349_cold-alcohol-used-dna-tests_htmlixzz1MkbEmEdu
httplearngeneticsutaheducontentlabsextractionhowto www kamibudaksainsblogspotcom
httpwwwchemwatch goldcom References and Resources Adapted from Berry Full of DNA by Diane
Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
58
REFERENCES
Uses of DNA Extraction | eHowcom httpwwwehowcomabout_5344428_uses-dna-extractionhtmlixzz1MkWQB2TZ
Science 10 A Contextual Approach Heineman Queensland Science Projects Regan Spence Maggie Spenceley
httpenwikipediaorgwikiMolecular_biology httpwwwclinchemorgcgicontent
extract44102201 http
employeescsbsjueduhjakubowskiclassesch331dnaoldnastructurehtml
httpwwwgenomebccaeducationteachersclassroom-activitiesdna-extraction
39
THE WHYrsquoS CUTTING PROTEIN AND DNA
The DNA in the nucleus of the cell is moulded folded and protected by proteins
40
THE WHYrsquoSALCOHOL Alcohol is less dense than water so it floats
on top Look for clumps of white stringy stuff where the water and alcohol layers meet
DNA is not soluble in alcohol - other cell parts are
By adding alcohol DNA precipitates out of the solution and collect at the interface of the alcohol and soap layer
The colder the alcohol the less soluble the DNA will be in it
41
COLD ALCOHOL
DNA dissolves in water but precipitates in alcohol
Cold alcohol is used to separate DNA out of water-based solutions
This allows the DNA to be purified for subsequent genetic testing
Adding alcohol to a solution containing DNA is a simple way to obtain the pure DNA required and colder temperatures slow down enzymes that can break down DNA giving better extraction results
42
WHY IS DNA EXTRACTION IMPORTANT DNA extraction is an important molecular
biology procedure
By definition extraction is taking DNA out of any type of cell for the purpose of analysis
See Handouts for more information on the use of DNA extracted (extension only)
43
SUMMARIZEDNA EXTRACTING FROM KIWI FRUIT
44
BREAK DOWN CELL WALLS
Cells walls have to be destroyed to reach the DNA within cells
Extraction procedures to obtain pure DNA have to get rid of all the molecular and chemical components of the tissue from which the DNA is being extracted
First steps involve lysing or destroying the cell walls This can be done with a variety of chemical agents that are caustic to cell membranes but do not harm the DNA Cells can also be sonicated homogenized or ground up to destroy membranes
45
REMOVING Removing Lipids
Once the cell walls have been destroyed a detergent is added to get rid of the fats and oils that make up the cell membranes Detergents cause the fats and oils to dissolve into the solution
Remove ProteinsProteins and enzymes can be digested by
adding a protease to the solution Proteases break down proteins into small peptides and amino acids
46
PRECIPITATING AND PURIFYING Precipitate DNA
Adding cold alcohol to a solution will cause DNA to precipitate and aggregate The DNA can be collected by centrifuging the sample and pouring off the liquid layer The DNA should exist as a small pellet in the bottom of the centrifuge tube
Purify DNAThe DNA can be washed by re-suspending it
in a cold alcohol solution and re-centrifuging it several times to obtain a very pure DNA sample Typical alcohols used to precipitate DNA include ethanol and Isopropanol This process leaves a very pure sample for stringent DNA testing
47
PART B OBSERVING EXTRACTED DNA UNDER THE MICROSCOPE
48
PART B A CLOSER LOOK
8 Use a dropping pipette to carefully remove some of the threadlike substance from the top of your preparation
9 Place one or two drops onto the middle of a
microscope slide
10 Add two drops of methylene blue Wait 3 or 4 minutes to allow the methylene blue to be absorbed by the DNA
11 Carefully place a cover slip on the slide Gently press
a folded piece of paper towelling over the top of the prepared slide to soak up any excess liquid
12 Observe the DNA under low power then high power
49
DISCUSSIONANSWER IN GROUP RELATED WORKED 1 Write a detailed description of the material
floating at the top of the test tube after the alcohol was added
2 Describe the DNA as it appears under the
microscope under high power 3 Prepare a diagram of your DNA specimen 4 What was the reason for using methylene
blue
50
HOMEWORKCONSTRUCT A FLOW CHART DRAWING MIND MAP OF THE PROCESS YOU USED EXTRACTING THE DNA FROM KIWIrsquoS
Next to each step explain how the method you used was important (Hint What substances make up the membranes of cells and cell organelles
How do detergents work
What is the active ingredient in meat tenderiser
51
TIPS STEPS FOR FLOW CHART In order to release the DNA from the
nuclei of the pea cells you first separated the cells from one another
Then the cell and nuclear membranes needed to be ruptured to release the cell contents and the contents of the nucleus
Once removed from the nuclear membrane the DNA had to be untangled into the visible threadlike structures you ended up with
52
HOMEWORK EXTENSION GROUP QUESTIONS Where can DNA be found in the cell Discuss the action of the soap (detergent) on the cell
What is the purpose of the soap in this activity What was the purpose of the Sodium Chloride Include a
discussion of polarity and charged particles Why was the cold ethanol added to the soap and salt
mixture Describe the appearance of your final product Draw a diagram of DNA containing 5 sets of nucleotide
bases labelling the hydrogen bonds between the bases References and Resources Adapted from Berry Full
of DNA by Diane Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
53
HTTPWWWGENOMEBCCAEDUCATIONTEACHERSCLASSROOM-ACTIVITIESDNA-EXTRACTION
54
EXTRACHROMOSOMES EXIST IN PAIRS Because our chromosomes exist in pairs (and consequently we
have 2 alleles of each gene) we are a diploid species This is why our somatic cells are represented as 2n Our gametes (sperm and ova) on the other hand are haploid and are represented as n Other species may have different ploidy for example
triploid (3n) seedless watermelons tetaploid (4n) salmonidae fish pentaploid (5n) Kenai birch hexaploid (6n) some types of wheat kiwi fruit octaploid (8n) acipenser (a genus
of sturgeon fish strawberies) decaploid (10n) some strawberries dodecaploid (12n) some types of amphibians eg Xenopus
ruwenzoriensis
wwwcyberusca
55
YOUR EXTRACTED DNA UNDER THE MICROSCOPE LOOKED
LIKE
Collaboration-Brainstorming on what to look at under the
- DNA extraction- DNA is too small for even a microscope to see and after the extraction it just appears like a blob True or false
Could you any DNA strand
wwwemployeescsbsjuedu
56
HANDOUT ON DNA EXTRACTIONBACKGROUND EVERY DAY LIFE
EXTRA READING MATERIAL(EXTENSION )
DISCOVERING DNA DNA ON THE INSIDE USE OF DNA EXTRACTION INN EVERYDAY LIFE WHY IS DNA TESTING GOOD MOLECULAR BIOLOGY PCR-based diagnostics of genomic DNA
57
REFERENCES httpwwwsquidoocomhow-to-get-dna-from-a-kiwi-fruit Why Is DNA Testing Good | eHowcom
httpwwwehowcomabout_6684410_dna-testing-good_htmlixzz1MkbcIJs5 Why Is DNA Extraction Important | eHowcom
httpwwwehowcomlist_5839095_dna-extraction-important_htmlixzz1MkZDrqyY
Why Is Sodium Used in DNA Extraction | eHowcom httpwwwehowcomabout_6504902_sodium-used-dna-extraction_htmlixzz1MkaGWg57
Why Is Cold Alcohol Used in DNA Tests | eHowcom httpwwwehowcomabout_6399349_cold-alcohol-used-dna-tests_htmlixzz1MkbEmEdu
httplearngeneticsutaheducontentlabsextractionhowto www kamibudaksainsblogspotcom
httpwwwchemwatch goldcom References and Resources Adapted from Berry Full of DNA by Diane
Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
58
REFERENCES
Uses of DNA Extraction | eHowcom httpwwwehowcomabout_5344428_uses-dna-extractionhtmlixzz1MkWQB2TZ
Science 10 A Contextual Approach Heineman Queensland Science Projects Regan Spence Maggie Spenceley
httpenwikipediaorgwikiMolecular_biology httpwwwclinchemorgcgicontent
extract44102201 http
employeescsbsjueduhjakubowskiclassesch331dnaoldnastructurehtml
httpwwwgenomebccaeducationteachersclassroom-activitiesdna-extraction
40
THE WHYrsquoSALCOHOL Alcohol is less dense than water so it floats
on top Look for clumps of white stringy stuff where the water and alcohol layers meet
DNA is not soluble in alcohol - other cell parts are
By adding alcohol DNA precipitates out of the solution and collect at the interface of the alcohol and soap layer
The colder the alcohol the less soluble the DNA will be in it
41
COLD ALCOHOL
DNA dissolves in water but precipitates in alcohol
Cold alcohol is used to separate DNA out of water-based solutions
This allows the DNA to be purified for subsequent genetic testing
Adding alcohol to a solution containing DNA is a simple way to obtain the pure DNA required and colder temperatures slow down enzymes that can break down DNA giving better extraction results
42
WHY IS DNA EXTRACTION IMPORTANT DNA extraction is an important molecular
biology procedure
By definition extraction is taking DNA out of any type of cell for the purpose of analysis
See Handouts for more information on the use of DNA extracted (extension only)
43
SUMMARIZEDNA EXTRACTING FROM KIWI FRUIT
44
BREAK DOWN CELL WALLS
Cells walls have to be destroyed to reach the DNA within cells
Extraction procedures to obtain pure DNA have to get rid of all the molecular and chemical components of the tissue from which the DNA is being extracted
First steps involve lysing or destroying the cell walls This can be done with a variety of chemical agents that are caustic to cell membranes but do not harm the DNA Cells can also be sonicated homogenized or ground up to destroy membranes
45
REMOVING Removing Lipids
Once the cell walls have been destroyed a detergent is added to get rid of the fats and oils that make up the cell membranes Detergents cause the fats and oils to dissolve into the solution
Remove ProteinsProteins and enzymes can be digested by
adding a protease to the solution Proteases break down proteins into small peptides and amino acids
46
PRECIPITATING AND PURIFYING Precipitate DNA
Adding cold alcohol to a solution will cause DNA to precipitate and aggregate The DNA can be collected by centrifuging the sample and pouring off the liquid layer The DNA should exist as a small pellet in the bottom of the centrifuge tube
Purify DNAThe DNA can be washed by re-suspending it
in a cold alcohol solution and re-centrifuging it several times to obtain a very pure DNA sample Typical alcohols used to precipitate DNA include ethanol and Isopropanol This process leaves a very pure sample for stringent DNA testing
47
PART B OBSERVING EXTRACTED DNA UNDER THE MICROSCOPE
48
PART B A CLOSER LOOK
8 Use a dropping pipette to carefully remove some of the threadlike substance from the top of your preparation
9 Place one or two drops onto the middle of a
microscope slide
10 Add two drops of methylene blue Wait 3 or 4 minutes to allow the methylene blue to be absorbed by the DNA
11 Carefully place a cover slip on the slide Gently press
a folded piece of paper towelling over the top of the prepared slide to soak up any excess liquid
12 Observe the DNA under low power then high power
49
DISCUSSIONANSWER IN GROUP RELATED WORKED 1 Write a detailed description of the material
floating at the top of the test tube after the alcohol was added
2 Describe the DNA as it appears under the
microscope under high power 3 Prepare a diagram of your DNA specimen 4 What was the reason for using methylene
blue
50
HOMEWORKCONSTRUCT A FLOW CHART DRAWING MIND MAP OF THE PROCESS YOU USED EXTRACTING THE DNA FROM KIWIrsquoS
Next to each step explain how the method you used was important (Hint What substances make up the membranes of cells and cell organelles
How do detergents work
What is the active ingredient in meat tenderiser
51
TIPS STEPS FOR FLOW CHART In order to release the DNA from the
nuclei of the pea cells you first separated the cells from one another
Then the cell and nuclear membranes needed to be ruptured to release the cell contents and the contents of the nucleus
Once removed from the nuclear membrane the DNA had to be untangled into the visible threadlike structures you ended up with
52
HOMEWORK EXTENSION GROUP QUESTIONS Where can DNA be found in the cell Discuss the action of the soap (detergent) on the cell
What is the purpose of the soap in this activity What was the purpose of the Sodium Chloride Include a
discussion of polarity and charged particles Why was the cold ethanol added to the soap and salt
mixture Describe the appearance of your final product Draw a diagram of DNA containing 5 sets of nucleotide
bases labelling the hydrogen bonds between the bases References and Resources Adapted from Berry Full
of DNA by Diane Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
53
HTTPWWWGENOMEBCCAEDUCATIONTEACHERSCLASSROOM-ACTIVITIESDNA-EXTRACTION
54
EXTRACHROMOSOMES EXIST IN PAIRS Because our chromosomes exist in pairs (and consequently we
have 2 alleles of each gene) we are a diploid species This is why our somatic cells are represented as 2n Our gametes (sperm and ova) on the other hand are haploid and are represented as n Other species may have different ploidy for example
triploid (3n) seedless watermelons tetaploid (4n) salmonidae fish pentaploid (5n) Kenai birch hexaploid (6n) some types of wheat kiwi fruit octaploid (8n) acipenser (a genus
of sturgeon fish strawberies) decaploid (10n) some strawberries dodecaploid (12n) some types of amphibians eg Xenopus
ruwenzoriensis
wwwcyberusca
55
YOUR EXTRACTED DNA UNDER THE MICROSCOPE LOOKED
LIKE
Collaboration-Brainstorming on what to look at under the
- DNA extraction- DNA is too small for even a microscope to see and after the extraction it just appears like a blob True or false
Could you any DNA strand
wwwemployeescsbsjuedu
56
HANDOUT ON DNA EXTRACTIONBACKGROUND EVERY DAY LIFE
EXTRA READING MATERIAL(EXTENSION )
DISCOVERING DNA DNA ON THE INSIDE USE OF DNA EXTRACTION INN EVERYDAY LIFE WHY IS DNA TESTING GOOD MOLECULAR BIOLOGY PCR-based diagnostics of genomic DNA
57
REFERENCES httpwwwsquidoocomhow-to-get-dna-from-a-kiwi-fruit Why Is DNA Testing Good | eHowcom
httpwwwehowcomabout_6684410_dna-testing-good_htmlixzz1MkbcIJs5 Why Is DNA Extraction Important | eHowcom
httpwwwehowcomlist_5839095_dna-extraction-important_htmlixzz1MkZDrqyY
Why Is Sodium Used in DNA Extraction | eHowcom httpwwwehowcomabout_6504902_sodium-used-dna-extraction_htmlixzz1MkaGWg57
Why Is Cold Alcohol Used in DNA Tests | eHowcom httpwwwehowcomabout_6399349_cold-alcohol-used-dna-tests_htmlixzz1MkbEmEdu
httplearngeneticsutaheducontentlabsextractionhowto www kamibudaksainsblogspotcom
httpwwwchemwatch goldcom References and Resources Adapted from Berry Full of DNA by Diane
Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
58
REFERENCES
Uses of DNA Extraction | eHowcom httpwwwehowcomabout_5344428_uses-dna-extractionhtmlixzz1MkWQB2TZ
Science 10 A Contextual Approach Heineman Queensland Science Projects Regan Spence Maggie Spenceley
httpenwikipediaorgwikiMolecular_biology httpwwwclinchemorgcgicontent
extract44102201 http
employeescsbsjueduhjakubowskiclassesch331dnaoldnastructurehtml
httpwwwgenomebccaeducationteachersclassroom-activitiesdna-extraction
41
COLD ALCOHOL
DNA dissolves in water but precipitates in alcohol
Cold alcohol is used to separate DNA out of water-based solutions
This allows the DNA to be purified for subsequent genetic testing
Adding alcohol to a solution containing DNA is a simple way to obtain the pure DNA required and colder temperatures slow down enzymes that can break down DNA giving better extraction results
42
WHY IS DNA EXTRACTION IMPORTANT DNA extraction is an important molecular
biology procedure
By definition extraction is taking DNA out of any type of cell for the purpose of analysis
See Handouts for more information on the use of DNA extracted (extension only)
43
SUMMARIZEDNA EXTRACTING FROM KIWI FRUIT
44
BREAK DOWN CELL WALLS
Cells walls have to be destroyed to reach the DNA within cells
Extraction procedures to obtain pure DNA have to get rid of all the molecular and chemical components of the tissue from which the DNA is being extracted
First steps involve lysing or destroying the cell walls This can be done with a variety of chemical agents that are caustic to cell membranes but do not harm the DNA Cells can also be sonicated homogenized or ground up to destroy membranes
45
REMOVING Removing Lipids
Once the cell walls have been destroyed a detergent is added to get rid of the fats and oils that make up the cell membranes Detergents cause the fats and oils to dissolve into the solution
Remove ProteinsProteins and enzymes can be digested by
adding a protease to the solution Proteases break down proteins into small peptides and amino acids
46
PRECIPITATING AND PURIFYING Precipitate DNA
Adding cold alcohol to a solution will cause DNA to precipitate and aggregate The DNA can be collected by centrifuging the sample and pouring off the liquid layer The DNA should exist as a small pellet in the bottom of the centrifuge tube
Purify DNAThe DNA can be washed by re-suspending it
in a cold alcohol solution and re-centrifuging it several times to obtain a very pure DNA sample Typical alcohols used to precipitate DNA include ethanol and Isopropanol This process leaves a very pure sample for stringent DNA testing
47
PART B OBSERVING EXTRACTED DNA UNDER THE MICROSCOPE
48
PART B A CLOSER LOOK
8 Use a dropping pipette to carefully remove some of the threadlike substance from the top of your preparation
9 Place one or two drops onto the middle of a
microscope slide
10 Add two drops of methylene blue Wait 3 or 4 minutes to allow the methylene blue to be absorbed by the DNA
11 Carefully place a cover slip on the slide Gently press
a folded piece of paper towelling over the top of the prepared slide to soak up any excess liquid
12 Observe the DNA under low power then high power
49
DISCUSSIONANSWER IN GROUP RELATED WORKED 1 Write a detailed description of the material
floating at the top of the test tube after the alcohol was added
2 Describe the DNA as it appears under the
microscope under high power 3 Prepare a diagram of your DNA specimen 4 What was the reason for using methylene
blue
50
HOMEWORKCONSTRUCT A FLOW CHART DRAWING MIND MAP OF THE PROCESS YOU USED EXTRACTING THE DNA FROM KIWIrsquoS
Next to each step explain how the method you used was important (Hint What substances make up the membranes of cells and cell organelles
How do detergents work
What is the active ingredient in meat tenderiser
51
TIPS STEPS FOR FLOW CHART In order to release the DNA from the
nuclei of the pea cells you first separated the cells from one another
Then the cell and nuclear membranes needed to be ruptured to release the cell contents and the contents of the nucleus
Once removed from the nuclear membrane the DNA had to be untangled into the visible threadlike structures you ended up with
52
HOMEWORK EXTENSION GROUP QUESTIONS Where can DNA be found in the cell Discuss the action of the soap (detergent) on the cell
What is the purpose of the soap in this activity What was the purpose of the Sodium Chloride Include a
discussion of polarity and charged particles Why was the cold ethanol added to the soap and salt
mixture Describe the appearance of your final product Draw a diagram of DNA containing 5 sets of nucleotide
bases labelling the hydrogen bonds between the bases References and Resources Adapted from Berry Full
of DNA by Diane Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
53
HTTPWWWGENOMEBCCAEDUCATIONTEACHERSCLASSROOM-ACTIVITIESDNA-EXTRACTION
54
EXTRACHROMOSOMES EXIST IN PAIRS Because our chromosomes exist in pairs (and consequently we
have 2 alleles of each gene) we are a diploid species This is why our somatic cells are represented as 2n Our gametes (sperm and ova) on the other hand are haploid and are represented as n Other species may have different ploidy for example
triploid (3n) seedless watermelons tetaploid (4n) salmonidae fish pentaploid (5n) Kenai birch hexaploid (6n) some types of wheat kiwi fruit octaploid (8n) acipenser (a genus
of sturgeon fish strawberies) decaploid (10n) some strawberries dodecaploid (12n) some types of amphibians eg Xenopus
ruwenzoriensis
wwwcyberusca
55
YOUR EXTRACTED DNA UNDER THE MICROSCOPE LOOKED
LIKE
Collaboration-Brainstorming on what to look at under the
- DNA extraction- DNA is too small for even a microscope to see and after the extraction it just appears like a blob True or false
Could you any DNA strand
wwwemployeescsbsjuedu
56
HANDOUT ON DNA EXTRACTIONBACKGROUND EVERY DAY LIFE
EXTRA READING MATERIAL(EXTENSION )
DISCOVERING DNA DNA ON THE INSIDE USE OF DNA EXTRACTION INN EVERYDAY LIFE WHY IS DNA TESTING GOOD MOLECULAR BIOLOGY PCR-based diagnostics of genomic DNA
57
REFERENCES httpwwwsquidoocomhow-to-get-dna-from-a-kiwi-fruit Why Is DNA Testing Good | eHowcom
httpwwwehowcomabout_6684410_dna-testing-good_htmlixzz1MkbcIJs5 Why Is DNA Extraction Important | eHowcom
httpwwwehowcomlist_5839095_dna-extraction-important_htmlixzz1MkZDrqyY
Why Is Sodium Used in DNA Extraction | eHowcom httpwwwehowcomabout_6504902_sodium-used-dna-extraction_htmlixzz1MkaGWg57
Why Is Cold Alcohol Used in DNA Tests | eHowcom httpwwwehowcomabout_6399349_cold-alcohol-used-dna-tests_htmlixzz1MkbEmEdu
httplearngeneticsutaheducontentlabsextractionhowto www kamibudaksainsblogspotcom
httpwwwchemwatch goldcom References and Resources Adapted from Berry Full of DNA by Diane
Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
58
REFERENCES
Uses of DNA Extraction | eHowcom httpwwwehowcomabout_5344428_uses-dna-extractionhtmlixzz1MkWQB2TZ
Science 10 A Contextual Approach Heineman Queensland Science Projects Regan Spence Maggie Spenceley
httpenwikipediaorgwikiMolecular_biology httpwwwclinchemorgcgicontent
extract44102201 http
employeescsbsjueduhjakubowskiclassesch331dnaoldnastructurehtml
httpwwwgenomebccaeducationteachersclassroom-activitiesdna-extraction
42
WHY IS DNA EXTRACTION IMPORTANT DNA extraction is an important molecular
biology procedure
By definition extraction is taking DNA out of any type of cell for the purpose of analysis
See Handouts for more information on the use of DNA extracted (extension only)
43
SUMMARIZEDNA EXTRACTING FROM KIWI FRUIT
44
BREAK DOWN CELL WALLS
Cells walls have to be destroyed to reach the DNA within cells
Extraction procedures to obtain pure DNA have to get rid of all the molecular and chemical components of the tissue from which the DNA is being extracted
First steps involve lysing or destroying the cell walls This can be done with a variety of chemical agents that are caustic to cell membranes but do not harm the DNA Cells can also be sonicated homogenized or ground up to destroy membranes
45
REMOVING Removing Lipids
Once the cell walls have been destroyed a detergent is added to get rid of the fats and oils that make up the cell membranes Detergents cause the fats and oils to dissolve into the solution
Remove ProteinsProteins and enzymes can be digested by
adding a protease to the solution Proteases break down proteins into small peptides and amino acids
46
PRECIPITATING AND PURIFYING Precipitate DNA
Adding cold alcohol to a solution will cause DNA to precipitate and aggregate The DNA can be collected by centrifuging the sample and pouring off the liquid layer The DNA should exist as a small pellet in the bottom of the centrifuge tube
Purify DNAThe DNA can be washed by re-suspending it
in a cold alcohol solution and re-centrifuging it several times to obtain a very pure DNA sample Typical alcohols used to precipitate DNA include ethanol and Isopropanol This process leaves a very pure sample for stringent DNA testing
47
PART B OBSERVING EXTRACTED DNA UNDER THE MICROSCOPE
48
PART B A CLOSER LOOK
8 Use a dropping pipette to carefully remove some of the threadlike substance from the top of your preparation
9 Place one or two drops onto the middle of a
microscope slide
10 Add two drops of methylene blue Wait 3 or 4 minutes to allow the methylene blue to be absorbed by the DNA
11 Carefully place a cover slip on the slide Gently press
a folded piece of paper towelling over the top of the prepared slide to soak up any excess liquid
12 Observe the DNA under low power then high power
49
DISCUSSIONANSWER IN GROUP RELATED WORKED 1 Write a detailed description of the material
floating at the top of the test tube after the alcohol was added
2 Describe the DNA as it appears under the
microscope under high power 3 Prepare a diagram of your DNA specimen 4 What was the reason for using methylene
blue
50
HOMEWORKCONSTRUCT A FLOW CHART DRAWING MIND MAP OF THE PROCESS YOU USED EXTRACTING THE DNA FROM KIWIrsquoS
Next to each step explain how the method you used was important (Hint What substances make up the membranes of cells and cell organelles
How do detergents work
What is the active ingredient in meat tenderiser
51
TIPS STEPS FOR FLOW CHART In order to release the DNA from the
nuclei of the pea cells you first separated the cells from one another
Then the cell and nuclear membranes needed to be ruptured to release the cell contents and the contents of the nucleus
Once removed from the nuclear membrane the DNA had to be untangled into the visible threadlike structures you ended up with
52
HOMEWORK EXTENSION GROUP QUESTIONS Where can DNA be found in the cell Discuss the action of the soap (detergent) on the cell
What is the purpose of the soap in this activity What was the purpose of the Sodium Chloride Include a
discussion of polarity and charged particles Why was the cold ethanol added to the soap and salt
mixture Describe the appearance of your final product Draw a diagram of DNA containing 5 sets of nucleotide
bases labelling the hydrogen bonds between the bases References and Resources Adapted from Berry Full
of DNA by Diane Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
53
HTTPWWWGENOMEBCCAEDUCATIONTEACHERSCLASSROOM-ACTIVITIESDNA-EXTRACTION
54
EXTRACHROMOSOMES EXIST IN PAIRS Because our chromosomes exist in pairs (and consequently we
have 2 alleles of each gene) we are a diploid species This is why our somatic cells are represented as 2n Our gametes (sperm and ova) on the other hand are haploid and are represented as n Other species may have different ploidy for example
triploid (3n) seedless watermelons tetaploid (4n) salmonidae fish pentaploid (5n) Kenai birch hexaploid (6n) some types of wheat kiwi fruit octaploid (8n) acipenser (a genus
of sturgeon fish strawberies) decaploid (10n) some strawberries dodecaploid (12n) some types of amphibians eg Xenopus
ruwenzoriensis
wwwcyberusca
55
YOUR EXTRACTED DNA UNDER THE MICROSCOPE LOOKED
LIKE
Collaboration-Brainstorming on what to look at under the
- DNA extraction- DNA is too small for even a microscope to see and after the extraction it just appears like a blob True or false
Could you any DNA strand
wwwemployeescsbsjuedu
56
HANDOUT ON DNA EXTRACTIONBACKGROUND EVERY DAY LIFE
EXTRA READING MATERIAL(EXTENSION )
DISCOVERING DNA DNA ON THE INSIDE USE OF DNA EXTRACTION INN EVERYDAY LIFE WHY IS DNA TESTING GOOD MOLECULAR BIOLOGY PCR-based diagnostics of genomic DNA
57
REFERENCES httpwwwsquidoocomhow-to-get-dna-from-a-kiwi-fruit Why Is DNA Testing Good | eHowcom
httpwwwehowcomabout_6684410_dna-testing-good_htmlixzz1MkbcIJs5 Why Is DNA Extraction Important | eHowcom
httpwwwehowcomlist_5839095_dna-extraction-important_htmlixzz1MkZDrqyY
Why Is Sodium Used in DNA Extraction | eHowcom httpwwwehowcomabout_6504902_sodium-used-dna-extraction_htmlixzz1MkaGWg57
Why Is Cold Alcohol Used in DNA Tests | eHowcom httpwwwehowcomabout_6399349_cold-alcohol-used-dna-tests_htmlixzz1MkbEmEdu
httplearngeneticsutaheducontentlabsextractionhowto www kamibudaksainsblogspotcom
httpwwwchemwatch goldcom References and Resources Adapted from Berry Full of DNA by Diane
Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
58
REFERENCES
Uses of DNA Extraction | eHowcom httpwwwehowcomabout_5344428_uses-dna-extractionhtmlixzz1MkWQB2TZ
Science 10 A Contextual Approach Heineman Queensland Science Projects Regan Spence Maggie Spenceley
httpenwikipediaorgwikiMolecular_biology httpwwwclinchemorgcgicontent
extract44102201 http
employeescsbsjueduhjakubowskiclassesch331dnaoldnastructurehtml
httpwwwgenomebccaeducationteachersclassroom-activitiesdna-extraction
43
SUMMARIZEDNA EXTRACTING FROM KIWI FRUIT
44
BREAK DOWN CELL WALLS
Cells walls have to be destroyed to reach the DNA within cells
Extraction procedures to obtain pure DNA have to get rid of all the molecular and chemical components of the tissue from which the DNA is being extracted
First steps involve lysing or destroying the cell walls This can be done with a variety of chemical agents that are caustic to cell membranes but do not harm the DNA Cells can also be sonicated homogenized or ground up to destroy membranes
45
REMOVING Removing Lipids
Once the cell walls have been destroyed a detergent is added to get rid of the fats and oils that make up the cell membranes Detergents cause the fats and oils to dissolve into the solution
Remove ProteinsProteins and enzymes can be digested by
adding a protease to the solution Proteases break down proteins into small peptides and amino acids
46
PRECIPITATING AND PURIFYING Precipitate DNA
Adding cold alcohol to a solution will cause DNA to precipitate and aggregate The DNA can be collected by centrifuging the sample and pouring off the liquid layer The DNA should exist as a small pellet in the bottom of the centrifuge tube
Purify DNAThe DNA can be washed by re-suspending it
in a cold alcohol solution and re-centrifuging it several times to obtain a very pure DNA sample Typical alcohols used to precipitate DNA include ethanol and Isopropanol This process leaves a very pure sample for stringent DNA testing
47
PART B OBSERVING EXTRACTED DNA UNDER THE MICROSCOPE
48
PART B A CLOSER LOOK
8 Use a dropping pipette to carefully remove some of the threadlike substance from the top of your preparation
9 Place one or two drops onto the middle of a
microscope slide
10 Add two drops of methylene blue Wait 3 or 4 minutes to allow the methylene blue to be absorbed by the DNA
11 Carefully place a cover slip on the slide Gently press
a folded piece of paper towelling over the top of the prepared slide to soak up any excess liquid
12 Observe the DNA under low power then high power
49
DISCUSSIONANSWER IN GROUP RELATED WORKED 1 Write a detailed description of the material
floating at the top of the test tube after the alcohol was added
2 Describe the DNA as it appears under the
microscope under high power 3 Prepare a diagram of your DNA specimen 4 What was the reason for using methylene
blue
50
HOMEWORKCONSTRUCT A FLOW CHART DRAWING MIND MAP OF THE PROCESS YOU USED EXTRACTING THE DNA FROM KIWIrsquoS
Next to each step explain how the method you used was important (Hint What substances make up the membranes of cells and cell organelles
How do detergents work
What is the active ingredient in meat tenderiser
51
TIPS STEPS FOR FLOW CHART In order to release the DNA from the
nuclei of the pea cells you first separated the cells from one another
Then the cell and nuclear membranes needed to be ruptured to release the cell contents and the contents of the nucleus
Once removed from the nuclear membrane the DNA had to be untangled into the visible threadlike structures you ended up with
52
HOMEWORK EXTENSION GROUP QUESTIONS Where can DNA be found in the cell Discuss the action of the soap (detergent) on the cell
What is the purpose of the soap in this activity What was the purpose of the Sodium Chloride Include a
discussion of polarity and charged particles Why was the cold ethanol added to the soap and salt
mixture Describe the appearance of your final product Draw a diagram of DNA containing 5 sets of nucleotide
bases labelling the hydrogen bonds between the bases References and Resources Adapted from Berry Full
of DNA by Diane Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
53
HTTPWWWGENOMEBCCAEDUCATIONTEACHERSCLASSROOM-ACTIVITIESDNA-EXTRACTION
54
EXTRACHROMOSOMES EXIST IN PAIRS Because our chromosomes exist in pairs (and consequently we
have 2 alleles of each gene) we are a diploid species This is why our somatic cells are represented as 2n Our gametes (sperm and ova) on the other hand are haploid and are represented as n Other species may have different ploidy for example
triploid (3n) seedless watermelons tetaploid (4n) salmonidae fish pentaploid (5n) Kenai birch hexaploid (6n) some types of wheat kiwi fruit octaploid (8n) acipenser (a genus
of sturgeon fish strawberies) decaploid (10n) some strawberries dodecaploid (12n) some types of amphibians eg Xenopus
ruwenzoriensis
wwwcyberusca
55
YOUR EXTRACTED DNA UNDER THE MICROSCOPE LOOKED
LIKE
Collaboration-Brainstorming on what to look at under the
- DNA extraction- DNA is too small for even a microscope to see and after the extraction it just appears like a blob True or false
Could you any DNA strand
wwwemployeescsbsjuedu
56
HANDOUT ON DNA EXTRACTIONBACKGROUND EVERY DAY LIFE
EXTRA READING MATERIAL(EXTENSION )
DISCOVERING DNA DNA ON THE INSIDE USE OF DNA EXTRACTION INN EVERYDAY LIFE WHY IS DNA TESTING GOOD MOLECULAR BIOLOGY PCR-based diagnostics of genomic DNA
57
REFERENCES httpwwwsquidoocomhow-to-get-dna-from-a-kiwi-fruit Why Is DNA Testing Good | eHowcom
httpwwwehowcomabout_6684410_dna-testing-good_htmlixzz1MkbcIJs5 Why Is DNA Extraction Important | eHowcom
httpwwwehowcomlist_5839095_dna-extraction-important_htmlixzz1MkZDrqyY
Why Is Sodium Used in DNA Extraction | eHowcom httpwwwehowcomabout_6504902_sodium-used-dna-extraction_htmlixzz1MkaGWg57
Why Is Cold Alcohol Used in DNA Tests | eHowcom httpwwwehowcomabout_6399349_cold-alcohol-used-dna-tests_htmlixzz1MkbEmEdu
httplearngeneticsutaheducontentlabsextractionhowto www kamibudaksainsblogspotcom
httpwwwchemwatch goldcom References and Resources Adapted from Berry Full of DNA by Diane
Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
58
REFERENCES
Uses of DNA Extraction | eHowcom httpwwwehowcomabout_5344428_uses-dna-extractionhtmlixzz1MkWQB2TZ
Science 10 A Contextual Approach Heineman Queensland Science Projects Regan Spence Maggie Spenceley
httpenwikipediaorgwikiMolecular_biology httpwwwclinchemorgcgicontent
extract44102201 http
employeescsbsjueduhjakubowskiclassesch331dnaoldnastructurehtml
httpwwwgenomebccaeducationteachersclassroom-activitiesdna-extraction
44
BREAK DOWN CELL WALLS
Cells walls have to be destroyed to reach the DNA within cells
Extraction procedures to obtain pure DNA have to get rid of all the molecular and chemical components of the tissue from which the DNA is being extracted
First steps involve lysing or destroying the cell walls This can be done with a variety of chemical agents that are caustic to cell membranes but do not harm the DNA Cells can also be sonicated homogenized or ground up to destroy membranes
45
REMOVING Removing Lipids
Once the cell walls have been destroyed a detergent is added to get rid of the fats and oils that make up the cell membranes Detergents cause the fats and oils to dissolve into the solution
Remove ProteinsProteins and enzymes can be digested by
adding a protease to the solution Proteases break down proteins into small peptides and amino acids
46
PRECIPITATING AND PURIFYING Precipitate DNA
Adding cold alcohol to a solution will cause DNA to precipitate and aggregate The DNA can be collected by centrifuging the sample and pouring off the liquid layer The DNA should exist as a small pellet in the bottom of the centrifuge tube
Purify DNAThe DNA can be washed by re-suspending it
in a cold alcohol solution and re-centrifuging it several times to obtain a very pure DNA sample Typical alcohols used to precipitate DNA include ethanol and Isopropanol This process leaves a very pure sample for stringent DNA testing
47
PART B OBSERVING EXTRACTED DNA UNDER THE MICROSCOPE
48
PART B A CLOSER LOOK
8 Use a dropping pipette to carefully remove some of the threadlike substance from the top of your preparation
9 Place one or two drops onto the middle of a
microscope slide
10 Add two drops of methylene blue Wait 3 or 4 minutes to allow the methylene blue to be absorbed by the DNA
11 Carefully place a cover slip on the slide Gently press
a folded piece of paper towelling over the top of the prepared slide to soak up any excess liquid
12 Observe the DNA under low power then high power
49
DISCUSSIONANSWER IN GROUP RELATED WORKED 1 Write a detailed description of the material
floating at the top of the test tube after the alcohol was added
2 Describe the DNA as it appears under the
microscope under high power 3 Prepare a diagram of your DNA specimen 4 What was the reason for using methylene
blue
50
HOMEWORKCONSTRUCT A FLOW CHART DRAWING MIND MAP OF THE PROCESS YOU USED EXTRACTING THE DNA FROM KIWIrsquoS
Next to each step explain how the method you used was important (Hint What substances make up the membranes of cells and cell organelles
How do detergents work
What is the active ingredient in meat tenderiser
51
TIPS STEPS FOR FLOW CHART In order to release the DNA from the
nuclei of the pea cells you first separated the cells from one another
Then the cell and nuclear membranes needed to be ruptured to release the cell contents and the contents of the nucleus
Once removed from the nuclear membrane the DNA had to be untangled into the visible threadlike structures you ended up with
52
HOMEWORK EXTENSION GROUP QUESTIONS Where can DNA be found in the cell Discuss the action of the soap (detergent) on the cell
What is the purpose of the soap in this activity What was the purpose of the Sodium Chloride Include a
discussion of polarity and charged particles Why was the cold ethanol added to the soap and salt
mixture Describe the appearance of your final product Draw a diagram of DNA containing 5 sets of nucleotide
bases labelling the hydrogen bonds between the bases References and Resources Adapted from Berry Full
of DNA by Diane Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
53
HTTPWWWGENOMEBCCAEDUCATIONTEACHERSCLASSROOM-ACTIVITIESDNA-EXTRACTION
54
EXTRACHROMOSOMES EXIST IN PAIRS Because our chromosomes exist in pairs (and consequently we
have 2 alleles of each gene) we are a diploid species This is why our somatic cells are represented as 2n Our gametes (sperm and ova) on the other hand are haploid and are represented as n Other species may have different ploidy for example
triploid (3n) seedless watermelons tetaploid (4n) salmonidae fish pentaploid (5n) Kenai birch hexaploid (6n) some types of wheat kiwi fruit octaploid (8n) acipenser (a genus
of sturgeon fish strawberies) decaploid (10n) some strawberries dodecaploid (12n) some types of amphibians eg Xenopus
ruwenzoriensis
wwwcyberusca
55
YOUR EXTRACTED DNA UNDER THE MICROSCOPE LOOKED
LIKE
Collaboration-Brainstorming on what to look at under the
- DNA extraction- DNA is too small for even a microscope to see and after the extraction it just appears like a blob True or false
Could you any DNA strand
wwwemployeescsbsjuedu
56
HANDOUT ON DNA EXTRACTIONBACKGROUND EVERY DAY LIFE
EXTRA READING MATERIAL(EXTENSION )
DISCOVERING DNA DNA ON THE INSIDE USE OF DNA EXTRACTION INN EVERYDAY LIFE WHY IS DNA TESTING GOOD MOLECULAR BIOLOGY PCR-based diagnostics of genomic DNA
57
REFERENCES httpwwwsquidoocomhow-to-get-dna-from-a-kiwi-fruit Why Is DNA Testing Good | eHowcom
httpwwwehowcomabout_6684410_dna-testing-good_htmlixzz1MkbcIJs5 Why Is DNA Extraction Important | eHowcom
httpwwwehowcomlist_5839095_dna-extraction-important_htmlixzz1MkZDrqyY
Why Is Sodium Used in DNA Extraction | eHowcom httpwwwehowcomabout_6504902_sodium-used-dna-extraction_htmlixzz1MkaGWg57
Why Is Cold Alcohol Used in DNA Tests | eHowcom httpwwwehowcomabout_6399349_cold-alcohol-used-dna-tests_htmlixzz1MkbEmEdu
httplearngeneticsutaheducontentlabsextractionhowto www kamibudaksainsblogspotcom
httpwwwchemwatch goldcom References and Resources Adapted from Berry Full of DNA by Diane
Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
58
REFERENCES
Uses of DNA Extraction | eHowcom httpwwwehowcomabout_5344428_uses-dna-extractionhtmlixzz1MkWQB2TZ
Science 10 A Contextual Approach Heineman Queensland Science Projects Regan Spence Maggie Spenceley
httpenwikipediaorgwikiMolecular_biology httpwwwclinchemorgcgicontent
extract44102201 http
employeescsbsjueduhjakubowskiclassesch331dnaoldnastructurehtml
httpwwwgenomebccaeducationteachersclassroom-activitiesdna-extraction
45
REMOVING Removing Lipids
Once the cell walls have been destroyed a detergent is added to get rid of the fats and oils that make up the cell membranes Detergents cause the fats and oils to dissolve into the solution
Remove ProteinsProteins and enzymes can be digested by
adding a protease to the solution Proteases break down proteins into small peptides and amino acids
46
PRECIPITATING AND PURIFYING Precipitate DNA
Adding cold alcohol to a solution will cause DNA to precipitate and aggregate The DNA can be collected by centrifuging the sample and pouring off the liquid layer The DNA should exist as a small pellet in the bottom of the centrifuge tube
Purify DNAThe DNA can be washed by re-suspending it
in a cold alcohol solution and re-centrifuging it several times to obtain a very pure DNA sample Typical alcohols used to precipitate DNA include ethanol and Isopropanol This process leaves a very pure sample for stringent DNA testing
47
PART B OBSERVING EXTRACTED DNA UNDER THE MICROSCOPE
48
PART B A CLOSER LOOK
8 Use a dropping pipette to carefully remove some of the threadlike substance from the top of your preparation
9 Place one or two drops onto the middle of a
microscope slide
10 Add two drops of methylene blue Wait 3 or 4 minutes to allow the methylene blue to be absorbed by the DNA
11 Carefully place a cover slip on the slide Gently press
a folded piece of paper towelling over the top of the prepared slide to soak up any excess liquid
12 Observe the DNA under low power then high power
49
DISCUSSIONANSWER IN GROUP RELATED WORKED 1 Write a detailed description of the material
floating at the top of the test tube after the alcohol was added
2 Describe the DNA as it appears under the
microscope under high power 3 Prepare a diagram of your DNA specimen 4 What was the reason for using methylene
blue
50
HOMEWORKCONSTRUCT A FLOW CHART DRAWING MIND MAP OF THE PROCESS YOU USED EXTRACTING THE DNA FROM KIWIrsquoS
Next to each step explain how the method you used was important (Hint What substances make up the membranes of cells and cell organelles
How do detergents work
What is the active ingredient in meat tenderiser
51
TIPS STEPS FOR FLOW CHART In order to release the DNA from the
nuclei of the pea cells you first separated the cells from one another
Then the cell and nuclear membranes needed to be ruptured to release the cell contents and the contents of the nucleus
Once removed from the nuclear membrane the DNA had to be untangled into the visible threadlike structures you ended up with
52
HOMEWORK EXTENSION GROUP QUESTIONS Where can DNA be found in the cell Discuss the action of the soap (detergent) on the cell
What is the purpose of the soap in this activity What was the purpose of the Sodium Chloride Include a
discussion of polarity and charged particles Why was the cold ethanol added to the soap and salt
mixture Describe the appearance of your final product Draw a diagram of DNA containing 5 sets of nucleotide
bases labelling the hydrogen bonds between the bases References and Resources Adapted from Berry Full
of DNA by Diane Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
53
HTTPWWWGENOMEBCCAEDUCATIONTEACHERSCLASSROOM-ACTIVITIESDNA-EXTRACTION
54
EXTRACHROMOSOMES EXIST IN PAIRS Because our chromosomes exist in pairs (and consequently we
have 2 alleles of each gene) we are a diploid species This is why our somatic cells are represented as 2n Our gametes (sperm and ova) on the other hand are haploid and are represented as n Other species may have different ploidy for example
triploid (3n) seedless watermelons tetaploid (4n) salmonidae fish pentaploid (5n) Kenai birch hexaploid (6n) some types of wheat kiwi fruit octaploid (8n) acipenser (a genus
of sturgeon fish strawberies) decaploid (10n) some strawberries dodecaploid (12n) some types of amphibians eg Xenopus
ruwenzoriensis
wwwcyberusca
55
YOUR EXTRACTED DNA UNDER THE MICROSCOPE LOOKED
LIKE
Collaboration-Brainstorming on what to look at under the
- DNA extraction- DNA is too small for even a microscope to see and after the extraction it just appears like a blob True or false
Could you any DNA strand
wwwemployeescsbsjuedu
56
HANDOUT ON DNA EXTRACTIONBACKGROUND EVERY DAY LIFE
EXTRA READING MATERIAL(EXTENSION )
DISCOVERING DNA DNA ON THE INSIDE USE OF DNA EXTRACTION INN EVERYDAY LIFE WHY IS DNA TESTING GOOD MOLECULAR BIOLOGY PCR-based diagnostics of genomic DNA
57
REFERENCES httpwwwsquidoocomhow-to-get-dna-from-a-kiwi-fruit Why Is DNA Testing Good | eHowcom
httpwwwehowcomabout_6684410_dna-testing-good_htmlixzz1MkbcIJs5 Why Is DNA Extraction Important | eHowcom
httpwwwehowcomlist_5839095_dna-extraction-important_htmlixzz1MkZDrqyY
Why Is Sodium Used in DNA Extraction | eHowcom httpwwwehowcomabout_6504902_sodium-used-dna-extraction_htmlixzz1MkaGWg57
Why Is Cold Alcohol Used in DNA Tests | eHowcom httpwwwehowcomabout_6399349_cold-alcohol-used-dna-tests_htmlixzz1MkbEmEdu
httplearngeneticsutaheducontentlabsextractionhowto www kamibudaksainsblogspotcom
httpwwwchemwatch goldcom References and Resources Adapted from Berry Full of DNA by Diane
Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
58
REFERENCES
Uses of DNA Extraction | eHowcom httpwwwehowcomabout_5344428_uses-dna-extractionhtmlixzz1MkWQB2TZ
Science 10 A Contextual Approach Heineman Queensland Science Projects Regan Spence Maggie Spenceley
httpenwikipediaorgwikiMolecular_biology httpwwwclinchemorgcgicontent
extract44102201 http
employeescsbsjueduhjakubowskiclassesch331dnaoldnastructurehtml
httpwwwgenomebccaeducationteachersclassroom-activitiesdna-extraction
46
PRECIPITATING AND PURIFYING Precipitate DNA
Adding cold alcohol to a solution will cause DNA to precipitate and aggregate The DNA can be collected by centrifuging the sample and pouring off the liquid layer The DNA should exist as a small pellet in the bottom of the centrifuge tube
Purify DNAThe DNA can be washed by re-suspending it
in a cold alcohol solution and re-centrifuging it several times to obtain a very pure DNA sample Typical alcohols used to precipitate DNA include ethanol and Isopropanol This process leaves a very pure sample for stringent DNA testing
47
PART B OBSERVING EXTRACTED DNA UNDER THE MICROSCOPE
48
PART B A CLOSER LOOK
8 Use a dropping pipette to carefully remove some of the threadlike substance from the top of your preparation
9 Place one or two drops onto the middle of a
microscope slide
10 Add two drops of methylene blue Wait 3 or 4 minutes to allow the methylene blue to be absorbed by the DNA
11 Carefully place a cover slip on the slide Gently press
a folded piece of paper towelling over the top of the prepared slide to soak up any excess liquid
12 Observe the DNA under low power then high power
49
DISCUSSIONANSWER IN GROUP RELATED WORKED 1 Write a detailed description of the material
floating at the top of the test tube after the alcohol was added
2 Describe the DNA as it appears under the
microscope under high power 3 Prepare a diagram of your DNA specimen 4 What was the reason for using methylene
blue
50
HOMEWORKCONSTRUCT A FLOW CHART DRAWING MIND MAP OF THE PROCESS YOU USED EXTRACTING THE DNA FROM KIWIrsquoS
Next to each step explain how the method you used was important (Hint What substances make up the membranes of cells and cell organelles
How do detergents work
What is the active ingredient in meat tenderiser
51
TIPS STEPS FOR FLOW CHART In order to release the DNA from the
nuclei of the pea cells you first separated the cells from one another
Then the cell and nuclear membranes needed to be ruptured to release the cell contents and the contents of the nucleus
Once removed from the nuclear membrane the DNA had to be untangled into the visible threadlike structures you ended up with
52
HOMEWORK EXTENSION GROUP QUESTIONS Where can DNA be found in the cell Discuss the action of the soap (detergent) on the cell
What is the purpose of the soap in this activity What was the purpose of the Sodium Chloride Include a
discussion of polarity and charged particles Why was the cold ethanol added to the soap and salt
mixture Describe the appearance of your final product Draw a diagram of DNA containing 5 sets of nucleotide
bases labelling the hydrogen bonds between the bases References and Resources Adapted from Berry Full
of DNA by Diane Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
53
HTTPWWWGENOMEBCCAEDUCATIONTEACHERSCLASSROOM-ACTIVITIESDNA-EXTRACTION
54
EXTRACHROMOSOMES EXIST IN PAIRS Because our chromosomes exist in pairs (and consequently we
have 2 alleles of each gene) we are a diploid species This is why our somatic cells are represented as 2n Our gametes (sperm and ova) on the other hand are haploid and are represented as n Other species may have different ploidy for example
triploid (3n) seedless watermelons tetaploid (4n) salmonidae fish pentaploid (5n) Kenai birch hexaploid (6n) some types of wheat kiwi fruit octaploid (8n) acipenser (a genus
of sturgeon fish strawberies) decaploid (10n) some strawberries dodecaploid (12n) some types of amphibians eg Xenopus
ruwenzoriensis
wwwcyberusca
55
YOUR EXTRACTED DNA UNDER THE MICROSCOPE LOOKED
LIKE
Collaboration-Brainstorming on what to look at under the
- DNA extraction- DNA is too small for even a microscope to see and after the extraction it just appears like a blob True or false
Could you any DNA strand
wwwemployeescsbsjuedu
56
HANDOUT ON DNA EXTRACTIONBACKGROUND EVERY DAY LIFE
EXTRA READING MATERIAL(EXTENSION )
DISCOVERING DNA DNA ON THE INSIDE USE OF DNA EXTRACTION INN EVERYDAY LIFE WHY IS DNA TESTING GOOD MOLECULAR BIOLOGY PCR-based diagnostics of genomic DNA
57
REFERENCES httpwwwsquidoocomhow-to-get-dna-from-a-kiwi-fruit Why Is DNA Testing Good | eHowcom
httpwwwehowcomabout_6684410_dna-testing-good_htmlixzz1MkbcIJs5 Why Is DNA Extraction Important | eHowcom
httpwwwehowcomlist_5839095_dna-extraction-important_htmlixzz1MkZDrqyY
Why Is Sodium Used in DNA Extraction | eHowcom httpwwwehowcomabout_6504902_sodium-used-dna-extraction_htmlixzz1MkaGWg57
Why Is Cold Alcohol Used in DNA Tests | eHowcom httpwwwehowcomabout_6399349_cold-alcohol-used-dna-tests_htmlixzz1MkbEmEdu
httplearngeneticsutaheducontentlabsextractionhowto www kamibudaksainsblogspotcom
httpwwwchemwatch goldcom References and Resources Adapted from Berry Full of DNA by Diane
Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
58
REFERENCES
Uses of DNA Extraction | eHowcom httpwwwehowcomabout_5344428_uses-dna-extractionhtmlixzz1MkWQB2TZ
Science 10 A Contextual Approach Heineman Queensland Science Projects Regan Spence Maggie Spenceley
httpenwikipediaorgwikiMolecular_biology httpwwwclinchemorgcgicontent
extract44102201 http
employeescsbsjueduhjakubowskiclassesch331dnaoldnastructurehtml
httpwwwgenomebccaeducationteachersclassroom-activitiesdna-extraction
47
PART B OBSERVING EXTRACTED DNA UNDER THE MICROSCOPE
48
PART B A CLOSER LOOK
8 Use a dropping pipette to carefully remove some of the threadlike substance from the top of your preparation
9 Place one or two drops onto the middle of a
microscope slide
10 Add two drops of methylene blue Wait 3 or 4 minutes to allow the methylene blue to be absorbed by the DNA
11 Carefully place a cover slip on the slide Gently press
a folded piece of paper towelling over the top of the prepared slide to soak up any excess liquid
12 Observe the DNA under low power then high power
49
DISCUSSIONANSWER IN GROUP RELATED WORKED 1 Write a detailed description of the material
floating at the top of the test tube after the alcohol was added
2 Describe the DNA as it appears under the
microscope under high power 3 Prepare a diagram of your DNA specimen 4 What was the reason for using methylene
blue
50
HOMEWORKCONSTRUCT A FLOW CHART DRAWING MIND MAP OF THE PROCESS YOU USED EXTRACTING THE DNA FROM KIWIrsquoS
Next to each step explain how the method you used was important (Hint What substances make up the membranes of cells and cell organelles
How do detergents work
What is the active ingredient in meat tenderiser
51
TIPS STEPS FOR FLOW CHART In order to release the DNA from the
nuclei of the pea cells you first separated the cells from one another
Then the cell and nuclear membranes needed to be ruptured to release the cell contents and the contents of the nucleus
Once removed from the nuclear membrane the DNA had to be untangled into the visible threadlike structures you ended up with
52
HOMEWORK EXTENSION GROUP QUESTIONS Where can DNA be found in the cell Discuss the action of the soap (detergent) on the cell
What is the purpose of the soap in this activity What was the purpose of the Sodium Chloride Include a
discussion of polarity and charged particles Why was the cold ethanol added to the soap and salt
mixture Describe the appearance of your final product Draw a diagram of DNA containing 5 sets of nucleotide
bases labelling the hydrogen bonds between the bases References and Resources Adapted from Berry Full
of DNA by Diane Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
53
HTTPWWWGENOMEBCCAEDUCATIONTEACHERSCLASSROOM-ACTIVITIESDNA-EXTRACTION
54
EXTRACHROMOSOMES EXIST IN PAIRS Because our chromosomes exist in pairs (and consequently we
have 2 alleles of each gene) we are a diploid species This is why our somatic cells are represented as 2n Our gametes (sperm and ova) on the other hand are haploid and are represented as n Other species may have different ploidy for example
triploid (3n) seedless watermelons tetaploid (4n) salmonidae fish pentaploid (5n) Kenai birch hexaploid (6n) some types of wheat kiwi fruit octaploid (8n) acipenser (a genus
of sturgeon fish strawberies) decaploid (10n) some strawberries dodecaploid (12n) some types of amphibians eg Xenopus
ruwenzoriensis
wwwcyberusca
55
YOUR EXTRACTED DNA UNDER THE MICROSCOPE LOOKED
LIKE
Collaboration-Brainstorming on what to look at under the
- DNA extraction- DNA is too small for even a microscope to see and after the extraction it just appears like a blob True or false
Could you any DNA strand
wwwemployeescsbsjuedu
56
HANDOUT ON DNA EXTRACTIONBACKGROUND EVERY DAY LIFE
EXTRA READING MATERIAL(EXTENSION )
DISCOVERING DNA DNA ON THE INSIDE USE OF DNA EXTRACTION INN EVERYDAY LIFE WHY IS DNA TESTING GOOD MOLECULAR BIOLOGY PCR-based diagnostics of genomic DNA
57
REFERENCES httpwwwsquidoocomhow-to-get-dna-from-a-kiwi-fruit Why Is DNA Testing Good | eHowcom
httpwwwehowcomabout_6684410_dna-testing-good_htmlixzz1MkbcIJs5 Why Is DNA Extraction Important | eHowcom
httpwwwehowcomlist_5839095_dna-extraction-important_htmlixzz1MkZDrqyY
Why Is Sodium Used in DNA Extraction | eHowcom httpwwwehowcomabout_6504902_sodium-used-dna-extraction_htmlixzz1MkaGWg57
Why Is Cold Alcohol Used in DNA Tests | eHowcom httpwwwehowcomabout_6399349_cold-alcohol-used-dna-tests_htmlixzz1MkbEmEdu
httplearngeneticsutaheducontentlabsextractionhowto www kamibudaksainsblogspotcom
httpwwwchemwatch goldcom References and Resources Adapted from Berry Full of DNA by Diane
Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
58
REFERENCES
Uses of DNA Extraction | eHowcom httpwwwehowcomabout_5344428_uses-dna-extractionhtmlixzz1MkWQB2TZ
Science 10 A Contextual Approach Heineman Queensland Science Projects Regan Spence Maggie Spenceley
httpenwikipediaorgwikiMolecular_biology httpwwwclinchemorgcgicontent
extract44102201 http
employeescsbsjueduhjakubowskiclassesch331dnaoldnastructurehtml
httpwwwgenomebccaeducationteachersclassroom-activitiesdna-extraction
48
PART B A CLOSER LOOK
8 Use a dropping pipette to carefully remove some of the threadlike substance from the top of your preparation
9 Place one or two drops onto the middle of a
microscope slide
10 Add two drops of methylene blue Wait 3 or 4 minutes to allow the methylene blue to be absorbed by the DNA
11 Carefully place a cover slip on the slide Gently press
a folded piece of paper towelling over the top of the prepared slide to soak up any excess liquid
12 Observe the DNA under low power then high power
49
DISCUSSIONANSWER IN GROUP RELATED WORKED 1 Write a detailed description of the material
floating at the top of the test tube after the alcohol was added
2 Describe the DNA as it appears under the
microscope under high power 3 Prepare a diagram of your DNA specimen 4 What was the reason for using methylene
blue
50
HOMEWORKCONSTRUCT A FLOW CHART DRAWING MIND MAP OF THE PROCESS YOU USED EXTRACTING THE DNA FROM KIWIrsquoS
Next to each step explain how the method you used was important (Hint What substances make up the membranes of cells and cell organelles
How do detergents work
What is the active ingredient in meat tenderiser
51
TIPS STEPS FOR FLOW CHART In order to release the DNA from the
nuclei of the pea cells you first separated the cells from one another
Then the cell and nuclear membranes needed to be ruptured to release the cell contents and the contents of the nucleus
Once removed from the nuclear membrane the DNA had to be untangled into the visible threadlike structures you ended up with
52
HOMEWORK EXTENSION GROUP QUESTIONS Where can DNA be found in the cell Discuss the action of the soap (detergent) on the cell
What is the purpose of the soap in this activity What was the purpose of the Sodium Chloride Include a
discussion of polarity and charged particles Why was the cold ethanol added to the soap and salt
mixture Describe the appearance of your final product Draw a diagram of DNA containing 5 sets of nucleotide
bases labelling the hydrogen bonds between the bases References and Resources Adapted from Berry Full
of DNA by Diane Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
53
HTTPWWWGENOMEBCCAEDUCATIONTEACHERSCLASSROOM-ACTIVITIESDNA-EXTRACTION
54
EXTRACHROMOSOMES EXIST IN PAIRS Because our chromosomes exist in pairs (and consequently we
have 2 alleles of each gene) we are a diploid species This is why our somatic cells are represented as 2n Our gametes (sperm and ova) on the other hand are haploid and are represented as n Other species may have different ploidy for example
triploid (3n) seedless watermelons tetaploid (4n) salmonidae fish pentaploid (5n) Kenai birch hexaploid (6n) some types of wheat kiwi fruit octaploid (8n) acipenser (a genus
of sturgeon fish strawberies) decaploid (10n) some strawberries dodecaploid (12n) some types of amphibians eg Xenopus
ruwenzoriensis
wwwcyberusca
55
YOUR EXTRACTED DNA UNDER THE MICROSCOPE LOOKED
LIKE
Collaboration-Brainstorming on what to look at under the
- DNA extraction- DNA is too small for even a microscope to see and after the extraction it just appears like a blob True or false
Could you any DNA strand
wwwemployeescsbsjuedu
56
HANDOUT ON DNA EXTRACTIONBACKGROUND EVERY DAY LIFE
EXTRA READING MATERIAL(EXTENSION )
DISCOVERING DNA DNA ON THE INSIDE USE OF DNA EXTRACTION INN EVERYDAY LIFE WHY IS DNA TESTING GOOD MOLECULAR BIOLOGY PCR-based diagnostics of genomic DNA
57
REFERENCES httpwwwsquidoocomhow-to-get-dna-from-a-kiwi-fruit Why Is DNA Testing Good | eHowcom
httpwwwehowcomabout_6684410_dna-testing-good_htmlixzz1MkbcIJs5 Why Is DNA Extraction Important | eHowcom
httpwwwehowcomlist_5839095_dna-extraction-important_htmlixzz1MkZDrqyY
Why Is Sodium Used in DNA Extraction | eHowcom httpwwwehowcomabout_6504902_sodium-used-dna-extraction_htmlixzz1MkaGWg57
Why Is Cold Alcohol Used in DNA Tests | eHowcom httpwwwehowcomabout_6399349_cold-alcohol-used-dna-tests_htmlixzz1MkbEmEdu
httplearngeneticsutaheducontentlabsextractionhowto www kamibudaksainsblogspotcom
httpwwwchemwatch goldcom References and Resources Adapted from Berry Full of DNA by Diane
Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
58
REFERENCES
Uses of DNA Extraction | eHowcom httpwwwehowcomabout_5344428_uses-dna-extractionhtmlixzz1MkWQB2TZ
Science 10 A Contextual Approach Heineman Queensland Science Projects Regan Spence Maggie Spenceley
httpenwikipediaorgwikiMolecular_biology httpwwwclinchemorgcgicontent
extract44102201 http
employeescsbsjueduhjakubowskiclassesch331dnaoldnastructurehtml
httpwwwgenomebccaeducationteachersclassroom-activitiesdna-extraction
49
DISCUSSIONANSWER IN GROUP RELATED WORKED 1 Write a detailed description of the material
floating at the top of the test tube after the alcohol was added
2 Describe the DNA as it appears under the
microscope under high power 3 Prepare a diagram of your DNA specimen 4 What was the reason for using methylene
blue
50
HOMEWORKCONSTRUCT A FLOW CHART DRAWING MIND MAP OF THE PROCESS YOU USED EXTRACTING THE DNA FROM KIWIrsquoS
Next to each step explain how the method you used was important (Hint What substances make up the membranes of cells and cell organelles
How do detergents work
What is the active ingredient in meat tenderiser
51
TIPS STEPS FOR FLOW CHART In order to release the DNA from the
nuclei of the pea cells you first separated the cells from one another
Then the cell and nuclear membranes needed to be ruptured to release the cell contents and the contents of the nucleus
Once removed from the nuclear membrane the DNA had to be untangled into the visible threadlike structures you ended up with
52
HOMEWORK EXTENSION GROUP QUESTIONS Where can DNA be found in the cell Discuss the action of the soap (detergent) on the cell
What is the purpose of the soap in this activity What was the purpose of the Sodium Chloride Include a
discussion of polarity and charged particles Why was the cold ethanol added to the soap and salt
mixture Describe the appearance of your final product Draw a diagram of DNA containing 5 sets of nucleotide
bases labelling the hydrogen bonds between the bases References and Resources Adapted from Berry Full
of DNA by Diane Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
53
HTTPWWWGENOMEBCCAEDUCATIONTEACHERSCLASSROOM-ACTIVITIESDNA-EXTRACTION
54
EXTRACHROMOSOMES EXIST IN PAIRS Because our chromosomes exist in pairs (and consequently we
have 2 alleles of each gene) we are a diploid species This is why our somatic cells are represented as 2n Our gametes (sperm and ova) on the other hand are haploid and are represented as n Other species may have different ploidy for example
triploid (3n) seedless watermelons tetaploid (4n) salmonidae fish pentaploid (5n) Kenai birch hexaploid (6n) some types of wheat kiwi fruit octaploid (8n) acipenser (a genus
of sturgeon fish strawberies) decaploid (10n) some strawberries dodecaploid (12n) some types of amphibians eg Xenopus
ruwenzoriensis
wwwcyberusca
55
YOUR EXTRACTED DNA UNDER THE MICROSCOPE LOOKED
LIKE
Collaboration-Brainstorming on what to look at under the
- DNA extraction- DNA is too small for even a microscope to see and after the extraction it just appears like a blob True or false
Could you any DNA strand
wwwemployeescsbsjuedu
56
HANDOUT ON DNA EXTRACTIONBACKGROUND EVERY DAY LIFE
EXTRA READING MATERIAL(EXTENSION )
DISCOVERING DNA DNA ON THE INSIDE USE OF DNA EXTRACTION INN EVERYDAY LIFE WHY IS DNA TESTING GOOD MOLECULAR BIOLOGY PCR-based diagnostics of genomic DNA
57
REFERENCES httpwwwsquidoocomhow-to-get-dna-from-a-kiwi-fruit Why Is DNA Testing Good | eHowcom
httpwwwehowcomabout_6684410_dna-testing-good_htmlixzz1MkbcIJs5 Why Is DNA Extraction Important | eHowcom
httpwwwehowcomlist_5839095_dna-extraction-important_htmlixzz1MkZDrqyY
Why Is Sodium Used in DNA Extraction | eHowcom httpwwwehowcomabout_6504902_sodium-used-dna-extraction_htmlixzz1MkaGWg57
Why Is Cold Alcohol Used in DNA Tests | eHowcom httpwwwehowcomabout_6399349_cold-alcohol-used-dna-tests_htmlixzz1MkbEmEdu
httplearngeneticsutaheducontentlabsextractionhowto www kamibudaksainsblogspotcom
httpwwwchemwatch goldcom References and Resources Adapted from Berry Full of DNA by Diane
Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
58
REFERENCES
Uses of DNA Extraction | eHowcom httpwwwehowcomabout_5344428_uses-dna-extractionhtmlixzz1MkWQB2TZ
Science 10 A Contextual Approach Heineman Queensland Science Projects Regan Spence Maggie Spenceley
httpenwikipediaorgwikiMolecular_biology httpwwwclinchemorgcgicontent
extract44102201 http
employeescsbsjueduhjakubowskiclassesch331dnaoldnastructurehtml
httpwwwgenomebccaeducationteachersclassroom-activitiesdna-extraction
50
HOMEWORKCONSTRUCT A FLOW CHART DRAWING MIND MAP OF THE PROCESS YOU USED EXTRACTING THE DNA FROM KIWIrsquoS
Next to each step explain how the method you used was important (Hint What substances make up the membranes of cells and cell organelles
How do detergents work
What is the active ingredient in meat tenderiser
51
TIPS STEPS FOR FLOW CHART In order to release the DNA from the
nuclei of the pea cells you first separated the cells from one another
Then the cell and nuclear membranes needed to be ruptured to release the cell contents and the contents of the nucleus
Once removed from the nuclear membrane the DNA had to be untangled into the visible threadlike structures you ended up with
52
HOMEWORK EXTENSION GROUP QUESTIONS Where can DNA be found in the cell Discuss the action of the soap (detergent) on the cell
What is the purpose of the soap in this activity What was the purpose of the Sodium Chloride Include a
discussion of polarity and charged particles Why was the cold ethanol added to the soap and salt
mixture Describe the appearance of your final product Draw a diagram of DNA containing 5 sets of nucleotide
bases labelling the hydrogen bonds between the bases References and Resources Adapted from Berry Full
of DNA by Diane Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
53
HTTPWWWGENOMEBCCAEDUCATIONTEACHERSCLASSROOM-ACTIVITIESDNA-EXTRACTION
54
EXTRACHROMOSOMES EXIST IN PAIRS Because our chromosomes exist in pairs (and consequently we
have 2 alleles of each gene) we are a diploid species This is why our somatic cells are represented as 2n Our gametes (sperm and ova) on the other hand are haploid and are represented as n Other species may have different ploidy for example
triploid (3n) seedless watermelons tetaploid (4n) salmonidae fish pentaploid (5n) Kenai birch hexaploid (6n) some types of wheat kiwi fruit octaploid (8n) acipenser (a genus
of sturgeon fish strawberies) decaploid (10n) some strawberries dodecaploid (12n) some types of amphibians eg Xenopus
ruwenzoriensis
wwwcyberusca
55
YOUR EXTRACTED DNA UNDER THE MICROSCOPE LOOKED
LIKE
Collaboration-Brainstorming on what to look at under the
- DNA extraction- DNA is too small for even a microscope to see and after the extraction it just appears like a blob True or false
Could you any DNA strand
wwwemployeescsbsjuedu
56
HANDOUT ON DNA EXTRACTIONBACKGROUND EVERY DAY LIFE
EXTRA READING MATERIAL(EXTENSION )
DISCOVERING DNA DNA ON THE INSIDE USE OF DNA EXTRACTION INN EVERYDAY LIFE WHY IS DNA TESTING GOOD MOLECULAR BIOLOGY PCR-based diagnostics of genomic DNA
57
REFERENCES httpwwwsquidoocomhow-to-get-dna-from-a-kiwi-fruit Why Is DNA Testing Good | eHowcom
httpwwwehowcomabout_6684410_dna-testing-good_htmlixzz1MkbcIJs5 Why Is DNA Extraction Important | eHowcom
httpwwwehowcomlist_5839095_dna-extraction-important_htmlixzz1MkZDrqyY
Why Is Sodium Used in DNA Extraction | eHowcom httpwwwehowcomabout_6504902_sodium-used-dna-extraction_htmlixzz1MkaGWg57
Why Is Cold Alcohol Used in DNA Tests | eHowcom httpwwwehowcomabout_6399349_cold-alcohol-used-dna-tests_htmlixzz1MkbEmEdu
httplearngeneticsutaheducontentlabsextractionhowto www kamibudaksainsblogspotcom
httpwwwchemwatch goldcom References and Resources Adapted from Berry Full of DNA by Diane
Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
58
REFERENCES
Uses of DNA Extraction | eHowcom httpwwwehowcomabout_5344428_uses-dna-extractionhtmlixzz1MkWQB2TZ
Science 10 A Contextual Approach Heineman Queensland Science Projects Regan Spence Maggie Spenceley
httpenwikipediaorgwikiMolecular_biology httpwwwclinchemorgcgicontent
extract44102201 http
employeescsbsjueduhjakubowskiclassesch331dnaoldnastructurehtml
httpwwwgenomebccaeducationteachersclassroom-activitiesdna-extraction
51
TIPS STEPS FOR FLOW CHART In order to release the DNA from the
nuclei of the pea cells you first separated the cells from one another
Then the cell and nuclear membranes needed to be ruptured to release the cell contents and the contents of the nucleus
Once removed from the nuclear membrane the DNA had to be untangled into the visible threadlike structures you ended up with
52
HOMEWORK EXTENSION GROUP QUESTIONS Where can DNA be found in the cell Discuss the action of the soap (detergent) on the cell
What is the purpose of the soap in this activity What was the purpose of the Sodium Chloride Include a
discussion of polarity and charged particles Why was the cold ethanol added to the soap and salt
mixture Describe the appearance of your final product Draw a diagram of DNA containing 5 sets of nucleotide
bases labelling the hydrogen bonds between the bases References and Resources Adapted from Berry Full
of DNA by Diane Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
53
HTTPWWWGENOMEBCCAEDUCATIONTEACHERSCLASSROOM-ACTIVITIESDNA-EXTRACTION
54
EXTRACHROMOSOMES EXIST IN PAIRS Because our chromosomes exist in pairs (and consequently we
have 2 alleles of each gene) we are a diploid species This is why our somatic cells are represented as 2n Our gametes (sperm and ova) on the other hand are haploid and are represented as n Other species may have different ploidy for example
triploid (3n) seedless watermelons tetaploid (4n) salmonidae fish pentaploid (5n) Kenai birch hexaploid (6n) some types of wheat kiwi fruit octaploid (8n) acipenser (a genus
of sturgeon fish strawberies) decaploid (10n) some strawberries dodecaploid (12n) some types of amphibians eg Xenopus
ruwenzoriensis
wwwcyberusca
55
YOUR EXTRACTED DNA UNDER THE MICROSCOPE LOOKED
LIKE
Collaboration-Brainstorming on what to look at under the
- DNA extraction- DNA is too small for even a microscope to see and after the extraction it just appears like a blob True or false
Could you any DNA strand
wwwemployeescsbsjuedu
56
HANDOUT ON DNA EXTRACTIONBACKGROUND EVERY DAY LIFE
EXTRA READING MATERIAL(EXTENSION )
DISCOVERING DNA DNA ON THE INSIDE USE OF DNA EXTRACTION INN EVERYDAY LIFE WHY IS DNA TESTING GOOD MOLECULAR BIOLOGY PCR-based diagnostics of genomic DNA
57
REFERENCES httpwwwsquidoocomhow-to-get-dna-from-a-kiwi-fruit Why Is DNA Testing Good | eHowcom
httpwwwehowcomabout_6684410_dna-testing-good_htmlixzz1MkbcIJs5 Why Is DNA Extraction Important | eHowcom
httpwwwehowcomlist_5839095_dna-extraction-important_htmlixzz1MkZDrqyY
Why Is Sodium Used in DNA Extraction | eHowcom httpwwwehowcomabout_6504902_sodium-used-dna-extraction_htmlixzz1MkaGWg57
Why Is Cold Alcohol Used in DNA Tests | eHowcom httpwwwehowcomabout_6399349_cold-alcohol-used-dna-tests_htmlixzz1MkbEmEdu
httplearngeneticsutaheducontentlabsextractionhowto www kamibudaksainsblogspotcom
httpwwwchemwatch goldcom References and Resources Adapted from Berry Full of DNA by Diane
Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
58
REFERENCES
Uses of DNA Extraction | eHowcom httpwwwehowcomabout_5344428_uses-dna-extractionhtmlixzz1MkWQB2TZ
Science 10 A Contextual Approach Heineman Queensland Science Projects Regan Spence Maggie Spenceley
httpenwikipediaorgwikiMolecular_biology httpwwwclinchemorgcgicontent
extract44102201 http
employeescsbsjueduhjakubowskiclassesch331dnaoldnastructurehtml
httpwwwgenomebccaeducationteachersclassroom-activitiesdna-extraction
52
HOMEWORK EXTENSION GROUP QUESTIONS Where can DNA be found in the cell Discuss the action of the soap (detergent) on the cell
What is the purpose of the soap in this activity What was the purpose of the Sodium Chloride Include a
discussion of polarity and charged particles Why was the cold ethanol added to the soap and salt
mixture Describe the appearance of your final product Draw a diagram of DNA containing 5 sets of nucleotide
bases labelling the hydrogen bonds between the bases References and Resources Adapted from Berry Full
of DNA by Diane Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
53
HTTPWWWGENOMEBCCAEDUCATIONTEACHERSCLASSROOM-ACTIVITIESDNA-EXTRACTION
54
EXTRACHROMOSOMES EXIST IN PAIRS Because our chromosomes exist in pairs (and consequently we
have 2 alleles of each gene) we are a diploid species This is why our somatic cells are represented as 2n Our gametes (sperm and ova) on the other hand are haploid and are represented as n Other species may have different ploidy for example
triploid (3n) seedless watermelons tetaploid (4n) salmonidae fish pentaploid (5n) Kenai birch hexaploid (6n) some types of wheat kiwi fruit octaploid (8n) acipenser (a genus
of sturgeon fish strawberies) decaploid (10n) some strawberries dodecaploid (12n) some types of amphibians eg Xenopus
ruwenzoriensis
wwwcyberusca
55
YOUR EXTRACTED DNA UNDER THE MICROSCOPE LOOKED
LIKE
Collaboration-Brainstorming on what to look at under the
- DNA extraction- DNA is too small for even a microscope to see and after the extraction it just appears like a blob True or false
Could you any DNA strand
wwwemployeescsbsjuedu
56
HANDOUT ON DNA EXTRACTIONBACKGROUND EVERY DAY LIFE
EXTRA READING MATERIAL(EXTENSION )
DISCOVERING DNA DNA ON THE INSIDE USE OF DNA EXTRACTION INN EVERYDAY LIFE WHY IS DNA TESTING GOOD MOLECULAR BIOLOGY PCR-based diagnostics of genomic DNA
57
REFERENCES httpwwwsquidoocomhow-to-get-dna-from-a-kiwi-fruit Why Is DNA Testing Good | eHowcom
httpwwwehowcomabout_6684410_dna-testing-good_htmlixzz1MkbcIJs5 Why Is DNA Extraction Important | eHowcom
httpwwwehowcomlist_5839095_dna-extraction-important_htmlixzz1MkZDrqyY
Why Is Sodium Used in DNA Extraction | eHowcom httpwwwehowcomabout_6504902_sodium-used-dna-extraction_htmlixzz1MkaGWg57
Why Is Cold Alcohol Used in DNA Tests | eHowcom httpwwwehowcomabout_6399349_cold-alcohol-used-dna-tests_htmlixzz1MkbEmEdu
httplearngeneticsutaheducontentlabsextractionhowto www kamibudaksainsblogspotcom
httpwwwchemwatch goldcom References and Resources Adapted from Berry Full of DNA by Diane
Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
58
REFERENCES
Uses of DNA Extraction | eHowcom httpwwwehowcomabout_5344428_uses-dna-extractionhtmlixzz1MkWQB2TZ
Science 10 A Contextual Approach Heineman Queensland Science Projects Regan Spence Maggie Spenceley
httpenwikipediaorgwikiMolecular_biology httpwwwclinchemorgcgicontent
extract44102201 http
employeescsbsjueduhjakubowskiclassesch331dnaoldnastructurehtml
httpwwwgenomebccaeducationteachersclassroom-activitiesdna-extraction
53
HTTPWWWGENOMEBCCAEDUCATIONTEACHERSCLASSROOM-ACTIVITIESDNA-EXTRACTION
54
EXTRACHROMOSOMES EXIST IN PAIRS Because our chromosomes exist in pairs (and consequently we
have 2 alleles of each gene) we are a diploid species This is why our somatic cells are represented as 2n Our gametes (sperm and ova) on the other hand are haploid and are represented as n Other species may have different ploidy for example
triploid (3n) seedless watermelons tetaploid (4n) salmonidae fish pentaploid (5n) Kenai birch hexaploid (6n) some types of wheat kiwi fruit octaploid (8n) acipenser (a genus
of sturgeon fish strawberies) decaploid (10n) some strawberries dodecaploid (12n) some types of amphibians eg Xenopus
ruwenzoriensis
wwwcyberusca
55
YOUR EXTRACTED DNA UNDER THE MICROSCOPE LOOKED
LIKE
Collaboration-Brainstorming on what to look at under the
- DNA extraction- DNA is too small for even a microscope to see and after the extraction it just appears like a blob True or false
Could you any DNA strand
wwwemployeescsbsjuedu
56
HANDOUT ON DNA EXTRACTIONBACKGROUND EVERY DAY LIFE
EXTRA READING MATERIAL(EXTENSION )
DISCOVERING DNA DNA ON THE INSIDE USE OF DNA EXTRACTION INN EVERYDAY LIFE WHY IS DNA TESTING GOOD MOLECULAR BIOLOGY PCR-based diagnostics of genomic DNA
57
REFERENCES httpwwwsquidoocomhow-to-get-dna-from-a-kiwi-fruit Why Is DNA Testing Good | eHowcom
httpwwwehowcomabout_6684410_dna-testing-good_htmlixzz1MkbcIJs5 Why Is DNA Extraction Important | eHowcom
httpwwwehowcomlist_5839095_dna-extraction-important_htmlixzz1MkZDrqyY
Why Is Sodium Used in DNA Extraction | eHowcom httpwwwehowcomabout_6504902_sodium-used-dna-extraction_htmlixzz1MkaGWg57
Why Is Cold Alcohol Used in DNA Tests | eHowcom httpwwwehowcomabout_6399349_cold-alcohol-used-dna-tests_htmlixzz1MkbEmEdu
httplearngeneticsutaheducontentlabsextractionhowto www kamibudaksainsblogspotcom
httpwwwchemwatch goldcom References and Resources Adapted from Berry Full of DNA by Diane
Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
58
REFERENCES
Uses of DNA Extraction | eHowcom httpwwwehowcomabout_5344428_uses-dna-extractionhtmlixzz1MkWQB2TZ
Science 10 A Contextual Approach Heineman Queensland Science Projects Regan Spence Maggie Spenceley
httpenwikipediaorgwikiMolecular_biology httpwwwclinchemorgcgicontent
extract44102201 http
employeescsbsjueduhjakubowskiclassesch331dnaoldnastructurehtml
httpwwwgenomebccaeducationteachersclassroom-activitiesdna-extraction
54
EXTRACHROMOSOMES EXIST IN PAIRS Because our chromosomes exist in pairs (and consequently we
have 2 alleles of each gene) we are a diploid species This is why our somatic cells are represented as 2n Our gametes (sperm and ova) on the other hand are haploid and are represented as n Other species may have different ploidy for example
triploid (3n) seedless watermelons tetaploid (4n) salmonidae fish pentaploid (5n) Kenai birch hexaploid (6n) some types of wheat kiwi fruit octaploid (8n) acipenser (a genus
of sturgeon fish strawberies) decaploid (10n) some strawberries dodecaploid (12n) some types of amphibians eg Xenopus
ruwenzoriensis
wwwcyberusca
55
YOUR EXTRACTED DNA UNDER THE MICROSCOPE LOOKED
LIKE
Collaboration-Brainstorming on what to look at under the
- DNA extraction- DNA is too small for even a microscope to see and after the extraction it just appears like a blob True or false
Could you any DNA strand
wwwemployeescsbsjuedu
56
HANDOUT ON DNA EXTRACTIONBACKGROUND EVERY DAY LIFE
EXTRA READING MATERIAL(EXTENSION )
DISCOVERING DNA DNA ON THE INSIDE USE OF DNA EXTRACTION INN EVERYDAY LIFE WHY IS DNA TESTING GOOD MOLECULAR BIOLOGY PCR-based diagnostics of genomic DNA
57
REFERENCES httpwwwsquidoocomhow-to-get-dna-from-a-kiwi-fruit Why Is DNA Testing Good | eHowcom
httpwwwehowcomabout_6684410_dna-testing-good_htmlixzz1MkbcIJs5 Why Is DNA Extraction Important | eHowcom
httpwwwehowcomlist_5839095_dna-extraction-important_htmlixzz1MkZDrqyY
Why Is Sodium Used in DNA Extraction | eHowcom httpwwwehowcomabout_6504902_sodium-used-dna-extraction_htmlixzz1MkaGWg57
Why Is Cold Alcohol Used in DNA Tests | eHowcom httpwwwehowcomabout_6399349_cold-alcohol-used-dna-tests_htmlixzz1MkbEmEdu
httplearngeneticsutaheducontentlabsextractionhowto www kamibudaksainsblogspotcom
httpwwwchemwatch goldcom References and Resources Adapted from Berry Full of DNA by Diane
Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
58
REFERENCES
Uses of DNA Extraction | eHowcom httpwwwehowcomabout_5344428_uses-dna-extractionhtmlixzz1MkWQB2TZ
Science 10 A Contextual Approach Heineman Queensland Science Projects Regan Spence Maggie Spenceley
httpenwikipediaorgwikiMolecular_biology httpwwwclinchemorgcgicontent
extract44102201 http
employeescsbsjueduhjakubowskiclassesch331dnaoldnastructurehtml
httpwwwgenomebccaeducationteachersclassroom-activitiesdna-extraction
55
YOUR EXTRACTED DNA UNDER THE MICROSCOPE LOOKED
LIKE
Collaboration-Brainstorming on what to look at under the
- DNA extraction- DNA is too small for even a microscope to see and after the extraction it just appears like a blob True or false
Could you any DNA strand
wwwemployeescsbsjuedu
56
HANDOUT ON DNA EXTRACTIONBACKGROUND EVERY DAY LIFE
EXTRA READING MATERIAL(EXTENSION )
DISCOVERING DNA DNA ON THE INSIDE USE OF DNA EXTRACTION INN EVERYDAY LIFE WHY IS DNA TESTING GOOD MOLECULAR BIOLOGY PCR-based diagnostics of genomic DNA
57
REFERENCES httpwwwsquidoocomhow-to-get-dna-from-a-kiwi-fruit Why Is DNA Testing Good | eHowcom
httpwwwehowcomabout_6684410_dna-testing-good_htmlixzz1MkbcIJs5 Why Is DNA Extraction Important | eHowcom
httpwwwehowcomlist_5839095_dna-extraction-important_htmlixzz1MkZDrqyY
Why Is Sodium Used in DNA Extraction | eHowcom httpwwwehowcomabout_6504902_sodium-used-dna-extraction_htmlixzz1MkaGWg57
Why Is Cold Alcohol Used in DNA Tests | eHowcom httpwwwehowcomabout_6399349_cold-alcohol-used-dna-tests_htmlixzz1MkbEmEdu
httplearngeneticsutaheducontentlabsextractionhowto www kamibudaksainsblogspotcom
httpwwwchemwatch goldcom References and Resources Adapted from Berry Full of DNA by Diane
Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
58
REFERENCES
Uses of DNA Extraction | eHowcom httpwwwehowcomabout_5344428_uses-dna-extractionhtmlixzz1MkWQB2TZ
Science 10 A Contextual Approach Heineman Queensland Science Projects Regan Spence Maggie Spenceley
httpenwikipediaorgwikiMolecular_biology httpwwwclinchemorgcgicontent
extract44102201 http
employeescsbsjueduhjakubowskiclassesch331dnaoldnastructurehtml
httpwwwgenomebccaeducationteachersclassroom-activitiesdna-extraction
56
HANDOUT ON DNA EXTRACTIONBACKGROUND EVERY DAY LIFE
EXTRA READING MATERIAL(EXTENSION )
DISCOVERING DNA DNA ON THE INSIDE USE OF DNA EXTRACTION INN EVERYDAY LIFE WHY IS DNA TESTING GOOD MOLECULAR BIOLOGY PCR-based diagnostics of genomic DNA
57
REFERENCES httpwwwsquidoocomhow-to-get-dna-from-a-kiwi-fruit Why Is DNA Testing Good | eHowcom
httpwwwehowcomabout_6684410_dna-testing-good_htmlixzz1MkbcIJs5 Why Is DNA Extraction Important | eHowcom
httpwwwehowcomlist_5839095_dna-extraction-important_htmlixzz1MkZDrqyY
Why Is Sodium Used in DNA Extraction | eHowcom httpwwwehowcomabout_6504902_sodium-used-dna-extraction_htmlixzz1MkaGWg57
Why Is Cold Alcohol Used in DNA Tests | eHowcom httpwwwehowcomabout_6399349_cold-alcohol-used-dna-tests_htmlixzz1MkbEmEdu
httplearngeneticsutaheducontentlabsextractionhowto www kamibudaksainsblogspotcom
httpwwwchemwatch goldcom References and Resources Adapted from Berry Full of DNA by Diane
Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
58
REFERENCES
Uses of DNA Extraction | eHowcom httpwwwehowcomabout_5344428_uses-dna-extractionhtmlixzz1MkWQB2TZ
Science 10 A Contextual Approach Heineman Queensland Science Projects Regan Spence Maggie Spenceley
httpenwikipediaorgwikiMolecular_biology httpwwwclinchemorgcgicontent
extract44102201 http
employeescsbsjueduhjakubowskiclassesch331dnaoldnastructurehtml
httpwwwgenomebccaeducationteachersclassroom-activitiesdna-extraction
57
REFERENCES httpwwwsquidoocomhow-to-get-dna-from-a-kiwi-fruit Why Is DNA Testing Good | eHowcom
httpwwwehowcomabout_6684410_dna-testing-good_htmlixzz1MkbcIJs5 Why Is DNA Extraction Important | eHowcom
httpwwwehowcomlist_5839095_dna-extraction-important_htmlixzz1MkZDrqyY
Why Is Sodium Used in DNA Extraction | eHowcom httpwwwehowcomabout_6504902_sodium-used-dna-extraction_htmlixzz1MkaGWg57
Why Is Cold Alcohol Used in DNA Tests | eHowcom httpwwwehowcomabout_6399349_cold-alcohol-used-dna-tests_htmlixzz1MkbEmEdu
httplearngeneticsutaheducontentlabsextractionhowto www kamibudaksainsblogspotcom
httpwwwchemwatch goldcom References and Resources Adapted from Berry Full of DNA by Diane
Sweeney for Biology Exploring Life to be published by Prentice Hall Websites httpwwwcarlinvilleschoolsnetlinkeBiologyDNAhtm httpcarnegieinstitutionorgfirst_light_casehornDNAdnaindexhtml
58
REFERENCES
Uses of DNA Extraction | eHowcom httpwwwehowcomabout_5344428_uses-dna-extractionhtmlixzz1MkWQB2TZ
Science 10 A Contextual Approach Heineman Queensland Science Projects Regan Spence Maggie Spenceley
httpenwikipediaorgwikiMolecular_biology httpwwwclinchemorgcgicontent
extract44102201 http
employeescsbsjueduhjakubowskiclassesch331dnaoldnastructurehtml
httpwwwgenomebccaeducationteachersclassroom-activitiesdna-extraction
58
REFERENCES
Uses of DNA Extraction | eHowcom httpwwwehowcomabout_5344428_uses-dna-extractionhtmlixzz1MkWQB2TZ
Science 10 A Contextual Approach Heineman Queensland Science Projects Regan Spence Maggie Spenceley
httpenwikipediaorgwikiMolecular_biology httpwwwclinchemorgcgicontent
extract44102201 http
employeescsbsjueduhjakubowskiclassesch331dnaoldnastructurehtml
httpwwwgenomebccaeducationteachersclassroom-activitiesdna-extraction
