DNA, RNA and Protein DNA, RNA and Protein Synthesis= CH 10Synthesis= CH 10

Griffith’s ExperimentsGriffith’s Experiments• Showed that hereditary material can pass from Showed that hereditary material can pass from

one bacterial cell to anotherone bacterial cell to another• The transfer of genetic material from one cell to The transfer of genetic material from one cell to

another or organism to organism is called another or organism to organism is called transformationtransformation

• Heat killed virulent bacteria can transfer their Heat killed virulent bacteria can transfer their disease causing ability to harmless bacteriadisease causing ability to harmless bacteria

Griffith’s ExperimentsGriffith’s Experiments

Avery’s ExperimentsAvery’s Experiments• Showed that: DNA is the hereditary Showed that: DNA is the hereditary

material that transfers info btwn material that transfers info btwn bacterial cellsbacterial cells

• Cells missing RNA and Protein could Cells missing RNA and Protein could transformtransform R into S cells R into S cells

• Cells missing DNA could not transform Cells missing DNA could not transform cellscells

Hershey-Chase ExperimentHershey-Chase Experiment• DNA not protein is the DNA not protein is the

genetic materialgenetic material

• DNA of viruses enters DNA of viruses enters bacterial cells and this bacterial cells and this causes the bacterial causes the bacterial cell to produce more cell to produce more viruses containing DNAviruses containing DNA

• Protein doesn’t enter Protein doesn’t enter cellscells

Discovery Of StructureDiscovery Of Structure• 1953:1953: Watson and CrickWatson and Crick

put together a model put together a model of DNA of DNA using Franklin’s and Wilkins’s using Franklin’s and Wilkins’s DNA diffraction X-raysDNA diffraction X-rays

• DNA is composed of 2 strands made of 4 DNA is composed of 2 strands made of 4 kinds of kinds of nucleotidesnucleotides

• Each nucleotide consists of 3 parts:Each nucleotide consists of 3 parts:

– one 5-carbon one 5-carbon sugar sugar (deoxyribose)(deoxyribose)

– one one phosphatephosphate group, and group, and

– one of 4 basesone of 4 bases

• adenine (adenine (AA), guanine (), guanine (GG), thymine (), thymine (TT), ), cytosine (cytosine (CC).).

Molecular Structure of DNAMolecular Structure of DNA

• Sugar & Phosphate Sugar & Phosphate are “sides” of ladder are “sides” of ladder and Bases are the and Bases are the “rungs” & attach to “rungs” & attach to sugarssugars

Structure of a nucleotideStructure of a nucleotide

2 categories of DNA bases:2 categories of DNA bases:Purines vs Pyrimidines Purines vs Pyrimidines

PURINES = A, G = SMALL WORD, BIG BASES = 2 RINGS

= PuAGPYRIMIDINES = T, C = BIG WORD, SMALL BASES= 1 RING = PyTC

Purines vs PyrimidinesPurines vs Pyrimidines• ChargaffChargaff showed that showed that

– % of % of AA always = always = % of % of TT

– % of % of GG always = always = % of % of CC

• Purines always with pyrimidinesPurines always with pyrimidines– BIG BASE ALWAYS WITH SMALLBIG BASE ALWAYS WITH SMALL

DNA DNA StructureStructure

Complementary base Complementary base pairing rulespairing rules

• Base pairsBase pairs are formed are formed by by

hydrogen hydrogen bondingbonding

of of A with T A with T

(2 H bonds)(2 H bonds), , and and

G with C (3 G with C (3 H bonds)H bonds)

DNA ReplicationDNA Replication

DNA Replication = DNA Replication = inin

S phase of cell S phase of cell cyclecycle

• An enzyme An enzyme ((helicasehelicase) breaks ) breaks the H bonds the H bonds between base pairs between base pairs and unZIPS the and unZIPS the strands = strands = replication forkreplication fork

• Another enzyme Another enzyme ((DNA polymeraseDNA polymerase) ) attaches the attaches the complementary complementary base to the original base to the original DNA strand DNA strand

DNA DNA ReplicationReplication

• Results in DNA Results in DNA molecules that consist molecules that consist of one "of one "oldold" strand and " strand and one "one "newnew" strand " strand

• Known as Known as semi-semi-conservative replication conservative replication b/c it conserves the b/c it conserves the original strandoriginal strand).).

DNA DNA ReplicationReplication

• Changes = mutationChanges = mutation

• Proofreading & repair prevent Proofreading & repair prevent many errorsmany errors

• Unrepaired mutation can cause Unrepaired mutation can cause cancercancer

DNA Errors in ReplicationDNA Errors in Replication

Flow of Flow of Genetic Genetic Material:Material:

DNA → RNA → ProteinsDNA → RNA → Proteins

RNA StructureRNA Structure• RNA differs from DNA RNA differs from DNA

– RNA uses RNA uses ribose as the sugar ribose as the sugar not deoxyribose.not deoxyribose.

– RNA bases are RNA bases are A, G, C, and A, G, C, and uraciluracil ( (UU).).

• G-CG-C• A-A-UU

– Single StrandedSingle Stranded– Shorter than DNAShorter than DNA

– Can Leave the nucleusCan Leave the nucleus

3 Types of RNA3 Types of RNA

• rRNA - ribosomalrRNA - ribosomal

• mRNA - messengermRNA - messenger

• tRNA - transfertRNA - transfer

Messenger RNA (mRNA)Messenger RNA (mRNA)• Made from DNA in nucleus

using RNA Polymerase• Is the “Is the “BlueprintBlueprint" for a " for a

protein protein – CarriedCarried to ribosomes in to ribosomes in

cytoplasm after “stop” is cytoplasm after “stop” is reachedreached

• Carries message from Carries message from nucleus to cytosolnucleus to cytosol

Ribosomal RNA (rRNA)Ribosomal RNA (rRNA)

• rRNA + protein makes a ribosomerRNA + protein makes a ribosome

• Site where proteins are assembled Site where proteins are assembled in cytoplasmin cytoplasm

Transfer RNA (tRNA)Transfer RNA (tRNA)

• CarriesCarries correct correct AA to AA to ribosome/ ribosome/ mRNA mRNA complexcomplex

TranscriptionTranscription

• DNA → RNADNA → RNA– uses RNA Polymerase (binds at uses RNA Polymerase (binds at

“promoter” region)“promoter” region)

– Process similar to DNA replicationProcess similar to DNA replication

– Begins with a START codon and ends Begins with a START codon and ends with a STOP codonwith a STOP codon

• Makes rRNA, tRNA or mRNAMakes rRNA, tRNA or mRNA• Message is “transcribed” from DNA code to RNA codeMessage is “transcribed” from DNA code to RNA code

TranscriptionTranscription

Protein Synthesis: TranslationProtein Synthesis: Translation

• Making of protein at the rRNA using Making of protein at the rRNA using mRNA and tRNAmRNA and tRNA

• Each base triplet in mRNA is called a Each base triplet in mRNA is called a codoncodon

--specifies an amino acid to be specifies an amino acid to be included into a polypeptide chainincluded into a polypeptide chain

–Uses genetic code to determine amino Uses genetic code to determine amino acidacid

Genetic CodeGenetic Code

• Universal for all forms of life Universal for all forms of life – 6161 triplets specifying amino acids triplets specifying amino acids

– 33 “stop” codes “stop” codes

• StopStop codes = codes = UAA, UAG, UGAUAA, UAG, UGA

• StartStart Codon = Codon = AUG = methionineAUG = methionine

From From DNA DNA

to Proteinsto Proteins

CODON CHART..from mRNA

TranslationTranslation• RNA → PROTEINRNA → PROTEIN• mRNA leaves nucleus goes to ribosomemRNA leaves nucleus goes to ribosome• Begins when ribosome attaches to start Begins when ribosome attaches to start

codoncodon• tRNA gets specific amino acid (floating free in tRNA gets specific amino acid (floating free in

cytosol), anticodon matches codon of mRNA cytosol), anticodon matches codon of mRNA and A.A.and A.A.

• tRNA brings its AA to ribosome and attaches tRNA brings its AA to ribosome and attaches it to growing chain of AA (protein)it to growing chain of AA (protein)

• stops at “stop” codonstops at “stop” codon

Chapter 11Chapter 11Gene ExpressionGene Expression

TURN “ON” GENES to TURN “ON” GENES to REGULATE PROTEIN AND REGULATE PROTEIN AND

GENE EXPRESSIONGENE EXPRESSION

Role of Gene ExpressionRole of Gene Expression

• Activation of a gene that results in Activation of a gene that results in transcription and production of transcription and production of mRNAmRNA

• Only a fraction of a cell’s genes are Only a fraction of a cell’s genes are expressed at any one timeexpressed at any one time–You only express genes or make You only express genes or make

proteins when NEEDED!proteins when NEEDED!

Gene Expression in ProkaryotesGene Expression in Prokaryotes-Studies in 1960’s by French -Studies in 1960’s by French scientistsscientists

-Started with simple intestinal -Started with simple intestinal prokaryotic cell= prokaryotic cell= Escherichia coliEscherichia coli = = E. coliE. coli

• Bacteria adapt to changes in their surroundings by using proteins to turn groups of genes on and off in response to various environmental signals

• The DNA of Escherichia coli is sufficient to encode about 4000 proteins, but only a fraction of these are made at any one time. E. coli regulates the expression of many of its genes according to the food sources that are available to it

• - Scientists discovered how genes - Scientists discovered how genes in this bacteria metabolize lactose in this bacteria metabolize lactose when when presentpresent

• -lactose = disaccharide…needs -lactose = disaccharide…needs to be broken down into galactose to be broken down into galactose and glucoseand glucose

Gene Expression in ProkaryotesGene Expression in Prokaryotes

• When lactose is absent, When lactose is absent, E. coliE. coli will will not produce the protein…is not produce the protein…is repressedrepressed

• When lactose is present, When lactose is present, E. coliE. coli will will produces the 3 structural enzymesproduces the 3 structural enzymes–Meaning this will make the “protein” Meaning this will make the “protein”

or go through induction…..so it can or go through induction…..so it can break down lactose!break down lactose!

Gene Expression in ProkaryotesGene Expression in Prokaryotes

• http://www.phschool.com/science/biology_place/biocoach/lacoperon/genereg.html

• GREAT ANIMATION TO REVIEW AT HOME!GREAT ANIMATION TO REVIEW AT HOME!

Gene Expression in ProkaryotesGene Expression in Prokaryotes

• Operon: series of genes coding for Operon: series of genes coding for specific products = “lac” operonspecific products = “lac” operon

• Operon = structural genes + Operon = structural genes + promoter + operatorpromoter + operator

Gene Expression in ProkaryotesGene Expression in Prokaryotes

• Promoter: segment of DNA Promoter: segment of DNA recognized by RNA polymerase recognized by RNA polymerase which then starts transcriptionwhich then starts transcription

• Operator: segment of DNA that Operator: segment of DNA that acts as “switch” by controlling the acts as “switch” by controlling the access of RNA polymerase to access of RNA polymerase to promoterpromoter

Prokaryotic On & Off switchesProkaryotic On & Off switches• Transcription can be turned “on or off” Transcription can be turned “on or off”

depending on what the cell needsdepending on what the cell needs

• When turned “off” a When turned “off” a repressorrepressor protein protein is bound to DNA in front of the geneis bound to DNA in front of the gene

• To turn a gene “on” an To turn a gene “on” an inducerinducer (lactose) binds to the repressor, (lactose) binds to the repressor, causing it to fall off….then gene is causing it to fall off….then gene is expressedexpressed

RepressionRepression

ActivationActivation

Gene Expression in EukaryotesGene Expression in Eukaryotes• Have not found “operons” in Have not found “operons” in

eukaryoteseukaryotes• Genomes are larger & more complexGenomes are larger & more complex• Organized into introns and exonsOrganized into introns and exons

– Through removal of introns from pre- Through removal of introns from pre- mRNAmRNA

Controlling Transcription in Controlling Transcription in EukaryotesEukaryotes

Removal of Introns After Removal of Introns After TranscriptionTranscription

Eukaryotic Genes are made of Eukaryotic Genes are made of introns and exonsintrons and exons

• IntronsIntrons noncoding noncoding portions of the gene, portions of the gene, removed by enzymes removed by enzymes before mRNA leaves the before mRNA leaves the nucleus (pre-mRNA)nucleus (pre-mRNA)

• ExonsExons portions that will portions that will eventually be translated eventually be translated remain in the finished remain in the finished mRNA that leaves the mRNA that leaves the nucleus.nucleus.

Gene Expression in DevelopmentGene Expression in Development• Expressed GenesExpressed Genes: have been transcribed & : have been transcribed &

translatedtranslated• Cell DifferentiationCell Differentiation: Development of cells w/ : Development of cells w/

different functionsdifferent functions• MorphogenesisMorphogenesis: development of form in an : development of form in an

organismorganism• Homeotic genes (hox):Homeotic genes (hox): determine where determine where

anatomical structures anatomical structures (appendages) will develop (appendages) will develop & controls differentiation & controls differentiation in early developmentin early development

Gene Expression in DevelopmentGene Expression in Development• Homeobox Sequence:Homeobox Sequence:

– w/in homeotic genesw/in homeotic genes

– Sequence of DNA that regulates Sequence of DNA that regulates patterns of developmentpatterns of development

– Homeoboxes of Homeoboxes of

many diff eukaryoticmany diff eukaryotic

organisms appearorganisms appear

to be very similarto be very similar

Gene Expression & CancerGene Expression & Cancer• OncogeneOncogene: Gene that causes cancer: Gene that causes cancer• Proto-oncogeneProto-oncogene = normal gene, = normal gene,

regulates cell growth. May mutate into regulates cell growth. May mutate into oncogeneoncogene that may lead to cancer that may lead to cancer

• Tumor-supressor gene (3 types)Tumor-supressor gene (3 types): for : for protein that prevents uncontrolled cell protein that prevents uncontrolled cell division, mutation may stop this protein division, mutation may stop this protein production production

• Viruses may have oncogenes or trigger Viruses may have oncogenes or trigger them in another cellthem in another cell

CancerCancer• Continue to divide indefinitely, even if too Continue to divide indefinitely, even if too

tightly packed or detach from other cellstightly packed or detach from other cells• TumorTumor: uncontrolled, abnormal cell division: uncontrolled, abnormal cell division• benign tumorbenign tumor: does not migrate to other areas, : does not migrate to other areas,

usually harmlessusually harmless• malignant tumormalignant tumor: invade other healthy tissues : invade other healthy tissues

= cancer= cancer• metastasismetastasis: breaking away and spreading to : breaking away and spreading to

other body parts to form new tumorsother body parts to form new tumors

Causes of CancerCauses of Cancer• CarcinogenCarcinogen

– Chemicals in tobacco smoke, asbestos, UV Chemicals in tobacco smoke, asbestos, UV light from the sunlight from the sun

– Mutagen: causes a mutationMutagen: causes a mutation

Kinds of Malignant TumorsKinds of Malignant Tumors• CarcinomaCarcinoma: in skin & tissue lining : in skin & tissue lining

organsorgans

• SarcomaSarcoma: in bone & muscle tissue: in bone & muscle tissue

• LymphomaLymphoma: in tissues that form : in tissues that form blood blood

• LeukemiaLeukemia: uncontrolled production : uncontrolled production of white blood cellsof white blood cells

Causes of CancerCauses of Cancer• Mutations that change expression of genes Mutations that change expression of genes

coding for growth factor proteinscoding for growth factor proteins• Usually comes after exposure to carcinogen Usually comes after exposure to carcinogen

(tobacco, UV light etc.)(tobacco, UV light etc.)• usually need more than 1 mutation to get cancerusually need more than 1 mutation to get cancer

Genetic Engineering Genetic Engineering and Biotechnology = and Biotechnology =

Ch 13Ch 13

DNA Identification/fingerprintingDNA Identification/fingerprinting

• Gene = segment of DNA bases that Gene = segment of DNA bases that code for traits and proteinscode for traits and proteins

• Genetic engineering= use of genes Genetic engineering= use of genes to create or modify the genometo create or modify the genome

• DNA fingerprinting = The repeating DNA fingerprinting = The repeating sequences in noncoding DNA sequences in noncoding DNA (introns) vary between individuals & (introns) vary between individuals & thus be used to identify an individualthus be used to identify an individual

Steps in DNA identification Steps in DNA identification (fingerprinting)(fingerprinting)

• Gel electrophoresis: pieces are separated Gel electrophoresis: pieces are separated by size on a gel creating “bands” = by size on a gel creating “bands” = fingerprintfingerprint

• Everybody has different number and size of Everybody has different number and size of pieces because their DNA sequences are pieces because their DNA sequences are differentdifferent

• PCR = polymerase chain reaction = PCR = polymerase chain reaction = duplicate DNAduplicate DNA– cut “digest” DNA with restriction enzyme to get cut “digest” DNA with restriction enzyme to get

a bunch of piecesa bunch of pieces

Gel ElectrophoresisGel Electrophoresis• DNA fragments placed into “wells” in DNA fragments placed into “wells” in

gel agarosegel agarose• Electricity pulls on DNA fragments, Electricity pulls on DNA fragments,

DNA is “-” and thus goes toward “+” DNA is “-” and thus goes toward “+” sideside

• Fragments travel at Fragments travel at different ratesdifferent rates based based on on sizesize and ability to and ability to squeeze through squeeze through swiss-cheese-like swiss-cheese-like agaroseagarose

DNA FingerprintingDNA Fingerprinting

DNA FingerprintingDNA Fingerprinting

Polymerase Chain Reaction (PCR)Polymerase Chain Reaction (PCR)• Useful if you only have a little bit of DNA Useful if you only have a little bit of DNA

and need to make copies of it and need to make copies of it

• Crime scenes, genetic disorders in Crime scenes, genetic disorders in embryonic cells, study ancient DNA embryonic cells, study ancient DNA fragmentsfragments

Restriction Restriction EnzymesEnzymes

• Cuts DNA at specific Cuts DNA at specific base sequencebase sequence

• Produces Produces sticky sticky endsends

• Recombinant DNA = Recombinant DNA = Complementary Complementary sticky ends can be sticky ends can be fused together…is fused together…is recombinedrecombined

Restriction EnzymesRestriction Enzymes

Producing Restriction FragmentsProducing Restriction Fragments

• DNA ligase enzyme used to splice DNA ligase enzyme used to splice together cut plasmids and together cut plasmids and chromosome fragmentschromosome fragments

Producing & combining restriction Producing & combining restriction fragmentsfragments

• Making identical copies of cellsMaking identical copies of cells

• Can clone genes or organismsCan clone genes or organisms

• Cloning a Gene= making large quantities of a desired DNA piece …usually Cloning a Gene= making large quantities of a desired DNA piece …usually

insert into a vector (bacteria)insert into a vector (bacteria)

• Transfers gene between organismsTransfers gene between organisms

• Plasmids:Plasmids: circle of DNA in bacterium replicates independently of the circle of DNA in bacterium replicates independently of the

single main chromosomesingle main chromosome

Cloning Cloning

• Gene may be used to make bacteria Gene may be used to make bacteria

produce specific protein - insulin produce specific protein - insulin

productionproduction

Transplanting GenesTransplanting Genes

Stem CellsStem Cells• Stem cells have the ability toStem cells have the ability to

1.1. divide and renew themselvesdivide and renew themselves

2.2. remain undifferentiated in formremain undifferentiated in form

33. develop into a variety of specialized cell types. develop into a variety of specialized cell types

Genomic LibraryGenomic Library• Includes all pieces of genome that Includes all pieces of genome that

come from cutting with a particular come from cutting with a particular restriction enzymerestriction enzyme

• Can have multiple libraries for the Can have multiple libraries for the same organism - all cut with different same organism - all cut with different R.E.’sR.E.’s

Transgenic OrganismTransgenic Organism• The host that has received the The host that has received the

recombinant DNArecombinant DNA

• Organism produces the new protein Organism produces the new protein unless the gene gets “turned off”unless the gene gets “turned off”

• Keep gene “turned on” by splicing it in Keep gene “turned on” by splicing it in near a gene that is frequently near a gene that is frequently expressedexpressed

Human Genome ProjectHuman Genome Project

• Sequence entire human genomeSequence entire human genome• Began in 1990 - expected completion Began in 1990 - expected completion

was 2005, but it was completed in was 2005, but it was completed in 2000 2000

• Thought humans had 100,000 genes, Thought humans had 100,000 genes, but its fewer than 30,000but its fewer than 30,000

• We have the sequence of genes, but We have the sequence of genes, but don’t know what they all do yetdon’t know what they all do yet

• Use info for diagnosis, treatment, Use info for diagnosis, treatment, prevention of genetic disordersprevention of genetic disorders

Future of GenomicsFuture of Genomics

• BioinformaticsBioinformatics: Uses computers to : Uses computers to catalog & analyze genomescatalog & analyze genomes

• ProteomicsProteomics: studies the identities, : studies the identities, interactions, and abundances of an interactions, and abundances of an organisms proteinsorganisms proteins

• MicroarraysMicroarrays: two-dimensional : two-dimensional arrangement of cloned genes, useful to arrangement of cloned genes, useful to compare specific proteins such as those compare specific proteins such as those that cause cancerthat cause cancer

Medical ApplicationsMedical Applications• Gene Therapy: Used on individuals to Gene Therapy: Used on individuals to

insert normal genes (or repair damaged insert normal genes (or repair damaged DNA) into DNA) into body cellsbody cells to to cure diseasecure disease– Abnormal gene can Abnormal gene can

still be inheritedstill be inherited

• Used on Used on fertilized zygotes or embryosfertilized zygotes or embryos to insert normal genes for both to insert normal genes for both developing body AND sex cellsdeveloping body AND sex cells– GenomeGenome changed permanently changed permanently

Uses of DNA TechnologyUses of DNA Technology

• CloningCloning• Stem Cell ResearchStem Cell Research• Pharmaceutical ProductsPharmaceutical Products

– insulininsulin• VaccinesVaccines

– work because body recognizes proteins, can work because body recognizes proteins, can produce protein without introducing produce protein without introducing pathogenpathogen

Uses of DNA TechnologyUses of DNA Technology• Agricultural CropsAgricultural Crops

– disease resistancedisease resistance– herbicide herbicide

resistanceresistance– Improve nutritionImprove nutrition– require less require less

fertilizer fertilizer (incorporate (incorporate nitrogen fixing nitrogen fixing gene)gene)

Concerns of DNA TechnologyConcerns of DNA Technology

• Plants might produce toxins that Plants might produce toxins that could cause allergies in people who could cause allergies in people who consume themconsume them

Concerns of DNA Concerns of DNA TechnologyTechnology

• What if the plants What if the plants get into the “wild” - get into the “wild” - forming forming “superweeds”“superweeds”

• Do we really know Do we really know what we are doing what we are doing when we mix genes?when we mix genes?