Strawberry DNA

1. Strawberry DNA Plant Genomics

2. Genomics – The study of DNA • Plant chromosomal DNA • Chromosome number • Plant genes • Plant reproduction • Plant gene expression – the regulation of genes

3. Why Plant Genomics • Agricultural applications – Production of healthier crops- more nutritious( Genetic engineering of crop plants • Production of crops with disease resistance • Pharmacology - What novel genes do plants have to apply to human pharmacological research? Many contain anti- cancer compounds • Bioremediation – Plants removing pollutants from the environment

4. Genomic DNA • 30,000 genes • Satellite DNA – around the centromeres • Telomeric DNA – repetitive copies of TTAGGG • VNTR – variable number of tandem repeats- variable per species • Retrotransposons – remains of ancient retroviruses – are capable of replicating and making enzymes capable of jumping our of one position and finding a new position in DNA

5. Genomic DNA • Sines- short interspersed elements – 500 bp but are not translated • Lines – long interspersed elements – up to 7000 bases some are transcribed and translated • Transposons- move around in the DNA • ALUs – a special type of DNA – 300 bp – accounts for 11 % of the human genome

6. Contradictions to the Central Dogma • Retroviruses – Other RNA viruses • Transposons • Other elements in DNA - Alus • Prions ( proteins – Mad Cow) • Genes for t- RNA • Genes for Ribosomal RNAs • Spliceosomes and catalytic RNA’s • Enhancers and repressors • Promoters

7. Materials • Clean blue tube • Sharpie marker • Eppendorf holder( Microfuge tubes) • Loading dye ( green and yellow tube) • Tracking dye – white tube TD • Marker – white tube – M • Strawberry DNA- sample from extraction

8. Extraction of DNA I • Homogenize strawberries • Filter the stawberry homogenate • Place extract in Corning Tube • Add lysis mixture – detergent containing lauryl sulfate and salt, NaCl • Add papain mixture to denature proteins( DNAses) • Mix by rocking and rolling

9. Extraction of DNA II • Heat at 55oC. This speeds up degradation of proteins( 2 min) • Place in ice until cold • Add ice cold ethanol. Drip slowly down the side of the Corning tube making sure that the alcohol forms a layer on top of the juice. • This should form an interface between the two layers.

10. DNA Precipitate III • The DNA should precipitate and form a mass of slender,sticky strands • Remove the DNA from the Corning tube, being careful not to disturb the interface • The DNA should be placed in a microcentrifuge tube, • Centrifuge for 2 minutes

11. Pellet and Supernatant • Spin the tube with the DNA • It forms a pellet on the side of the microcentrifuge tube • Pour off the supernatant which is alcohol • The DNA needs to be resolubilized in Tris buffer • Mix the DNA from the pellet with Tris

12. Tris and DNA • The DNA must be in solution for electrophoresis. • Mix before using in the gel • Remove 25 ul of DNA and place in a microcentrifuge tube. • Add 3 ul of loading dye • Vortex to mix

13. Genomic DNA – Gel LanesLabel on your index card • 1-Tracking dye • 2- Marker – Lambda phage - HindII • 3 -8- Strawberry DNA samples( genomic DNA) Name on Ziplock Snack Bag for Gel

15. Loading gels • Run DNA from black to red • From the cathode to the anode • Remember DNA has a negative charge

16. Reminders about loading gel • Aspirate to first stop point • Deliver by dispensing into the well to the second stop point • Fill the well – being careful not to overfill • Run the gel on a voltage of 80 volts

17. Staining • Stain with Methylene blue. Fast Blast Stain. Gel should stain for at least one hour • Destain with Distilled water • Place gel in ziplock bag with water. • When gel is sufficeintly destained - the stained gel can be placed in a ziplock for storage in the refrigerator

18. Gel Observation • Observe gel on light box • Measure the distance that each band in the marker( standard) migrates • Measure any bands oar streaks of DNA you observe. • Use the DNA graphing program to make a semi-log graph to estimate the sizes of your DNA

19. DNA gel