DNA Precipitation

DNA Precipitation

Phenol (removes protein)

1. add equal volume of Phenol (= tris-saturated Phenol-Chloroform-Isoamyethanol)
2. vortex
3. spin 2 minutes at 12000 rpm 4°C
4. transfer supernatant to a fresh tube (avoid aspiration of the interlayer or organic phase)

Chloroform (removes phenol)

1. add equal volume of Chloroform
2. vortex
3. spin 2 minutes at 12000 rpm 4°C
4. transfer supernatant to a fresh tube (avoid aspiration of the interlayer or organic phase)

100% Ethanol (precipitates DNA)

1. add 0.1 volume 3 M sodium acetate
2. add 2.5 volumes 100 % Ethanol
3. vortex
4. precipitate at:
   • -20°C overnight (+++)
   • -80°C 1 h (++)
   • dry ice 15min (+)
5. spin 20 minutes at 12000 rpm 4°C
6. carefully pour out / aspirate supernatant (do not lose DNA-pellet)

70% Ethanol (washes out salt)

1. carefully add 1 mL cold 70% Ethanol (do not vortex)
2. spin 10 minutes at 12000 rpm 4°C
3. carefully pour out / aspirate supernatant (do not lose DNA-pellet)
4. air dry 10 minutes at room temperature (do not overdry, because DNA becomes hard to dissolve)
5. dissolve in:
   • 10 mM Tris pH 7.5 (+++)
   • TE-Buffer (++) - EDTA may inhibit downstream enzymatic reactions
   • dH₂ O (+) - freeze at -20°C because unbuffered DNA undergoes degradation