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Making DNA relevant and exciting in the high school classroom
By Anna Heyer and Rachel Zenuk
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What is BioME?• Is it like a biome?
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Bio and Me?
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Biology from molecules to evolution?
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What is NSF’s GK-12 initiative?• ~100 5-year training grants nationwide• Funds grad student partnerships with K-12 public school teachers• Goal: fellows acquire communication and teaching skills, while enriching STEM education in K-12 classrooms• Training a new generation of scientists• Strengthening partnerships between universities & local school districts
What is the focus of BioME?
• Focus: teaching the life sciences via hands-on experiences, using a framework of evolution & biodiversity
We are entering the fourth of five years
What’s the goal of a teacher-fellowpartnership?
• Partners work together to develop and use classroomteaching materials in the life sciences
• Partnerships are year-long, with the fellow in the classroom every week
• The fellow is a resource with scientific expertise• The fellow is not an aide, sub, or student teacher
Adriana Racolta:planting a gardenat Liberty Elementary
Aletris Neilis:dissecting apuma (!) atHermosaMontessori
Matt Herron: teachingmolecular techniquesat Tucson High
BioME tapsinto fellows’individualskills andinterests.
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Sweet Water Wetland Project 3rd grade class at White Elementary
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Liberty Elementary Garden Club
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2nd Grade Insect Projects
Summer Institute
• Getting acquainted with BioME philosophy & aims
• Getting partnerships underway via team time
• Introducing teaching resources
• Discussing traits of successful partnerships
BioME forges strong bonds and lasting partnerships.
Who runs BioME?
Principal InvestigatorsJudie BronsteinBarry RothStacey Forsyth
Graduate Coordinator Kathleen Walker
Educational EvaluatorMelissa PageMelvin Hall
Administration
K-12 CoordinatorMary Bouley
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Our Partnership
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The Manduca Project
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AP Science Field Trip
• Goal- to increase rigor and interest in AP science and chemistry
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San Pedro River
• Students collected insect larva, turtles, and other aquatic specimens
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Bob cats, pumas, and snakes...Oh My!
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A great trip, but most of the sleeping happened on the bus
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DNA Extraction
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DNA Extraction
• Purpose– to excite students and “hook” them in on the
first or second day of school– to teach basic procedures and classroom
management– to introduce the molecule we will be working
with all year
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Materials
• Small vials
• rubbing alcohol or ethanol 70% or higher
• toothpicks
• liquid dish soap in water (1:2 soap to water)
• 1% salt solution
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Procedures
1. Swish 15 ml of salt water for 1 minute.
2. Pour spit mixture into conical soap tube
3. Rock back and forth for 2 minutes.
4. Add 5-10mL of chilled ethanol to the tube.
5. PULL OUT THE DNA!
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Summarize the procedures in your own words….
1. Swish with salt water
2. Mix with soap3. Add EtOH4. Twirl with
collection stick5. Preserve DNA
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How could use DNA extraction in your classroom?
• Extensions: create DNA necklaces
• Questions???
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Microarray Lab
Today’s BIG Idea
• Over the last 30 years many defects in genes have been linked to cancer, each promising to be the magic in understanding and curing cancer.
• We now know cancer is a multistep process, and accumulation of mutations, or genetic aberrations, allows a cell to progress to tumor and malignancy.
Cancer is caused by genetic mutations
Additions
Deletions
Normal gene
Single base change
DNA
CT
A G C G A A C TAC
A G G C G C T AAC A C T
A G C T A A C TAC
A G A A C TAC
Cancer is caused by genetic mutations
Cancer cell division
Fourth orlater mutation
Third mutation
Second mutation
First mutation
Uncontrolled growth
Cell Suicide or Apoptosis
Cell damage—no repair
Normal cell division
Cancer involves MULTIPLE mutations
Malignant cells invade neighboring tissues, enter blood vessels, and metastasize to different sites
More mutations, more genetic instability, metastatic disease
Proto-oncogenes mutate to oncogenes
Mutations inactivate DNA repair genes
Cells proliferate
Mutation inactivates suppressor gene
Benign tumor cells grow only locally and cannot spread by invasion or metastasis
Time
Oncogenes
Mutated/damaged oncogene
Oncogenes accelerate cell growth and division
Cancer cell
Normal cell Normal genes regulate cell growth
Tumor Suppressor Genes
Normal genes prevent cancer
Remove or inactivate tumor suppressor genes
Mutated/inactivated tumor suppressor genes
Damage to both genes leads to cancer
Cancer cell
Normal cell
DNA Repair Genes
Cancer
No cancer
No DNA repair
Normal DNA repair
Base pair mismatch
T CATC
A GTCG
T CAGC
A GTCG
A GTG A GTAG
T CATCT CATC
MANY Genes are Implicated in Cancer!• Every cancer can be attributed to a different set of genetic
aberrations, and different genes are either expressed or not expressed.
• More than 100 different types of cancer can be found within specific organs!
This makes cancer treatment tricky…• Each caner has a different potential of being treated by current
therapies.
• For example, it has been shown cancer cells that lack p53 do not respond well to radiation therapy, and other non-malignant cells lacking p53 will progress to malignancy in response to radiation.
• Thus the treatment itself can cause more cancers!
NORMAL cell Cell suicide (Apoptosis)
p53 protein
Excessive DNA damage
Discussion Questions
• How do you determine the function of a gene?
• Which genetic aberrations have been implicated in cancer?
• What cellular functions are affected (turned ON or OFF) in cancer cells, and how might these affect normal cell development?
What is the best way to treat cancer?• Figure out which genes are mutated and which
genes are expressed or not expressed in the tissue.
• Gene expression of numerous genes can be looked at by a new technique called microarray analysis.
What is microarray analysis?
Microarray analysis uses cDNA to look at gene expression
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DNA
mRNA
cDNAprotein
Transcription
TranslationReverse Transcription
cDNA is labeled with fluorescent dyes
Microarray analysis shows us which genes are expressed in cancer
Materials
• Microarray slide -- this slide contains genes involved in cancer
• Disposable pipette• cDNA mixture solution -- these cDNAs
were made from normal breast tissue (attached to blue dye) and breast cancer tissues (attached to red dye).
• Wash solution• Color developing reagent
Microarray SlideSymbol Name FunctionPOL1 DNA Polymerase DNA replicationGAPdH Glyc.Ald.Phos.DeH-ase Kreb CycleHK1 Hexokinase 1 GlycolysisALDH1 AldDeHase Converts retinal to retinoic acid, overexpression confers cyclophos. resistance.GLUT1 Glucose Transporter 1 Transports glucose molecules into cells for energyACTG1 Actin, cytoplasmic Microtubule formation, cytoskeleton formationDNASE1 Deoxyribonuclease I Degrades DNARNASE4 Ribonuclease 4 Degrades RNATOP1 Topoisomerase I Aids in DNA supercoilingBRCA1 Breast cancer type 1 susceptibility protein Plays a role in DNA double-strand break repairPDGFR Platelet-derived Growth Factor Receptor Integral membrane receptor that binds PDGFCYP1A1 Cytochrome P450 1A1 Drug metabolismBCL2 B-cell lymphoma protein 2 Supresses apoptosisLIG1 DNA Ligase I DNA Ligation during replication/repairPOL1 DNA Polymerase Iota Synthesizes DNA on a template strandAPAF1 Apoptosis Protease Activating Factor 1 Tumor suppressor- Promotes apoptosis in damaged/ irregular cellsp53 p53 (tumor protein 53) Tumor supressor- induces growth arrest and/or apoptosisZNF84 Zinc Finger Protein 84 May play a role in transcription regulationMUC1 Transmembrane Mucin 1 Plays a role in cell adhesion, cell to cell interactionsG6PD Glu.6-phosp DeH-ase Metabolism, Provides pentose sugars for nucleic acid synth.TNF Tumor necrosis factor Cytokine, may induce tumor cell death. Deficiencies common in cancerADH4 Alcohol Dehydrogenase Alcohol processingDNMT1 DNA Methyltranferase I Modifies DNA to make it inaccessible thereby inhibiting transcriptionPOLR2A RNA Polymerase, subunit 2 RNA Polymerase synthesizes RNAMDM2 MDM2 Inhibits p53-induced arrest and cell deathMMP3 Matrix Metalloprotease 3 (Stromelysin) Degrades extracellular matrix that anchors cells in placeVEGF Vascular endothelial growth factor Growth factor that promotes formation of new blood vesselsACAT1 Acetoacetyl-CoA thiolase Ketone body metabolismMCR4 Melanocortin receptor Binds melanocortin; multiple downstream effectsPDK2 Pyruvate DeH-ase Kinase Phosphorylates/inhibits PDH complexGPB Glycerol Phosphatase Beta Inhibits glycogen phosphorylaseDUSP1 Dual-specificity protein 1 Dephosphorylates and "resets" MAPKPRL1 Protein Tyrosine Phosphatase Stops growth signal cascade from receptor tyrosine kinasesJUN Jun Component of AP-1 transcription factor- activates transcriptionFOS Fos Component of AP-1 transcription factor- activates transcriptionRASSF1 Ras-association domain, family 1 protein Inhibits cell cycle progression at the G1-S phase transitionRAS ras Small G-protein, signaling molecule in transcription activationSOS sos Tyrosine-kinase receptor signaling molecule, binds SH3 domainsEGFR Epithelial Growth Factor Receptor Binds EGF to promote epithelial cell growthCS Citrate Synthase Kreb Cycle enzymeAChE Acetylcholinesterase Degrades Ach to stop action potential in nervesCDKNA p21 Works with p53 to stop cell cycle progressionIL6 Interleukin 6 Cytokine, differentiation of b-cells, nerve cellsGSTP1 Glutathione S-transferase Helps to inactivate and eliminate some types of toxinsVEGFR Vascular Endothelial Growth Factor Rec. Binds VEGF, promotes growth of new vasculaturePLCG1 Phospholipase-C gamma Cleaves Phosphatidyl Inositol TriPhosphate into IP3 and DAG for signalingMYC c-Myc Proto-oncogene Activates transcription of growth-related genesRPS18 Ribosome Subunit 18S Ribosomes translate mRNA into proteinNAT1 n-acetyltransferase Modifies histones, MAPK Mitogen-activated Protein Kinase Signaling molecule and transcriptional activator
Procedure1. Place the slide onto the paper towel.
2. Add enough of the cDNA solution to the slide to completely cover it, but not spill off of the slide.
3. Let the cDNA hybridize with the microarray slide for 5 minutes.
4. After the 5 minute incubation of the microarray slide with cDNA, rinse off the excess cDNA with the microarray wash solution (in squeeze bottle).
5. Add color solution, again enough to cover the slide but not spill over the slide. This solution is toxic so take care to not get it on you, and wash off of skin immediately. Let the color solution set for 30 sec, then wash off excess with microarray wash solution.
6. Record you data.
Results
Symbol Name Function5. GLUT1 Glucose Transporter 1 Transports glucose molecules into cells for energy
10. BRCA1Breast cancer type 1 susceptibility protein Plays a role in DNA double-strand break repair
13. BCL2 B-cell lymphoma protein 2 Suppresses apoptosis
16. APAF1Apoptosis Protease Activating Factor 1
Tumor suppressor- Promotes apoptosis in damaged/ irregular cells
23. DNMT1 DNA Methyltranferase IModifies DNA to make it inaccessible thereby inhibiting transcription
26. MMP3Matrix Metalloprotease 3 (Stromelysin) Degrades extracellular matrix that anchors cells in place
27. VEGF Vascular endothelial growth factorGrowth factor that promotes formation of new blood vessels
33. PRL1 Protein Tyrosine PhosphataseStops growth signal cascade from receptor tyrosine kinases
36. RASSF1Ras-association domain, family 1 protein
Inhibits cell cycle progression at the G1-S phase transition
44. GSTP1 Glutathione S-transferase Helps to inactivate and eliminate some types of toxins47. MYC c-Myc Proto-oncogene Activates transcription of growth-related genes
Discussion
• Microarray analysis shows us which genes are expresses in normal cells vs. cancer cells.
– Why are some genes expressed in normal cells?
– Why are some genes expressed in cancer cells?
– Why are some genes expressed in both?
Discussion
• Genes expressed in normal cells only are likely tumor suppressors or DNA repair genes.
• Genes expressed in cancer cells only are likely oncogenes.
• Genes expressed in both are present in both conditions.
THANK YOU!
Questions?Contact the BIOTECH Project:
Dr. Nadja Anderson
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