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LIMA BEAN BACTERIA DNA EXTRACTION

Lana Hays

Materials

dry lima beans
Palmolive detergent
centrifuge
distilled water
centrifuge tube
fresh papaya juice
graduated cylinder (10ml)
non-iodized salt
granulated sugar
pipet
epsom salts
15 ml test tube
Bufferin (325mg)
test tube rack or 250 ml beaker
ice cold 95% ethanol
glass stirring rod

Solutions

   
Lima Bean Bacteria Suspension:
Place 1-2 handfuls of dry lima beans in a large jar and fill halfway to the top with distilled water. Cover and sit in a warm room for 2-3 days. Culturing longer than three days often results in more DNA but it usually shears. Pour through a strainer and keep the liquid for the extractions.

Prep buffer solution:
57 g granulated sugar
3 g epsom salts
1 buffered aspirin
add distilled water for a total volume of 500 ml

50% detergent solution:
20 ml detergent
20 ml distilled water

Salt solution:
29.2 g non-iodized salt
add distilled water for a total volume of 250 ml

Protocol

  1. Add 14 ml of the bacterial suspension to a centrifuge tube and spin in a balanced centrifuge for 5 minutes.
  2. Pour off the liquid (supernatant) and discard. You want to keep the pellet as this has your cells.
  3. Add 5 ml of prep buffer and resuspend your cells with a pipet.
  4. Add 1 ml 50% detergent solution.
  5. Add 1 ml papaya juice.
  6. Add 2 ml salt solution and shake for 2 minutes.
  7. Place the tube in the centrifuge and spin for 5 minutes. Make sure the centrifuge is balanced.
  8. Draw off 7 ml of the supernatant (liquid) as this has the DNA and place it in a clean test tube.
  9. Pour 7 ml of ice cold ethanol carefully down the side of the tube.
  10. Let the mixture sit undisturbed 2-3 minutes until the bubbling stops.
  11. The DNA will float in the alcohol. Swirl a glass rod at the interface of the two layers. You may see some tiny threads of DNA but are more likely to see fluffy, white sheared DNA.
Modified from: "Generic, All Purpose DNA Extraction from Meat Protocol" Judy Brown "Mammalian DNA Extraction" Theresa Knapp


Onion | Wheat Germ | Lima Bean Bacteria | Thymus | Yeast | Data Sheet and Questions | Design Your Own DNA Extraction Protocol | Teacher Information | References


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