Strawberry DNA

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Strawberry DNA

• Plant Genomics

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Genomics The study of DNA

• Plant chromosomal DNA
• Chromosome number
• Plant genes
• Plant reproduction
• Plant gene expression the regulation of genes

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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

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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

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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

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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 RNAs
• Enhancers and repressors
• Promoters

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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

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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

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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.

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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

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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

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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

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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
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Loading gels

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

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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

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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

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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

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DNA gel
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