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DNA Extraction from Cheek

Cells
Sample Preparation

Cheek cells are obtained by rinsing the mouth with 25mls of any commercial mouth wash soluti
available for about 30sec the first thing in the morning. It is important not to brush your teeth be
collecting the specimen. The mouth wash is then discarded into a sterile conical tube and sent to
laboratory.

A Salting Out Procedure for DNA extraction from Cheek Cells

1. Tube containing mouthwash is centrifuged at 4000 rpm for 15 minutes.

2. Supernatant is removed cautiously by means of a sterile pipette, without disturbing the
at the bottom.
3. Pellet is washed with 25ml TE buffer, by means of centrifigation at 4000rpm for 10 min
4. Repeat steps 2 and 3.
5. The pellet is resuspended in 1ml SE buffer containing 1μl proteinase K and 1% SDS.
6. Incubate at 370C overnight in a water bath.
7. Add 1ml SE buffer
8. Add 500μl 6M NaCl
9. Add 2.5ml chloroform
10. Mix for 60 min in an over-the-top rotator.
11. Vortex for 20 sec.
12. Centrifuge for 10 minutes at 2000rpm, and set the slowest breaking force
13. Transfer the supernatant to a clean tube.
 14. Add an equal volume of isopropan-2-ol to the supernatant to precipitate the DNA.
15. If DNA is visible go to step 16 and if not go to step 17.
16. If DNA is visible, spool out the DNA and place in microfuge tube filled with 1ml of
ethanol (go to step 18)
17. If DNA is NOT visible centrifuge the tube for 4 minutes at 11,000g and discar
isopropanol. Add 1ml of 75% ethanol.
18. Centrifuge for 4 minutes at 11,000g and discard the ethanol taking care not to lose the D
19. Add ethanol and repeat centrifugation 2 times. Discard ethanol after last centrifugation.
20. Evaporate ethanol by placing microfuge tube in oven (max 60ºC) for about 20-30 mins.
21. Add 100μl of TE buffer to rehydrate DNA, and mix overnight on an over the top rotator.

This method was evaluated at the DNA Laboratory, Medical School, Malta.

(Mr. Joseph Borg)

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