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Phenol/Chloroform Precipitation of DNA

Phenol/Chloroform Precipitation of DNA



1. Add an equal volume (equal to sample volume) of P/C to sample.
2. Mix (shake, don't vortex).
3. Take aqueous (upper) layer. (If dirty sample, repeat Ph/Chl step until interface is fairly clean).
4. Add equal volume chloroform, mix (shake, don't vortex).
5. Spin 3 min.
6. Take aqueous (upper) layer. (Optional: repeat Chloroform steps).
7. Add 1/10 volume 3M NaOAc (-> 0.3M), mix (shake).
8. Add 2 volumes ice-cold EtOH (100%), mix (shake).
9. Incubate on ice for 15 to 30 minutes. (Can store on ice or at -20°ree;C at this step).
10. Spin 10 minutes, 4°ree;C.
11. Remove supernatant, being careful not to disturb pellet (DNA).
12. Half-fill tube with 70% EtOH, spin at least 2 minutes at 4°ree;C. (Opt: Re-rinse and spin again).
13. Pipet off the sup, careful not to touch pellet.
14. Air-dry (~ 1 hour).
15. Redissolve in 10 mM Tris.

Shirley Reynolds
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