The general procedure for precipitating DNA and RNA is:
1. Add one-tenth volume of 3M NaOAc, pH 5.2* to the nucleic acid solution to be precipitated,
2. Add two volumes of cold 95% ethanol,
3. Place at -70°C for at least 30 min, or at -20°C overnight. or alternatively
1. Combine 95 ml of 100% ethanol with 4 ml of 3 M NaOAc (pH 4.8) and 1ml of sterile water. Mix by inversion and store at -20°C.
2. Add 2.5 volumes of cold ethanol/acetate solution to the nucleic acid solution to be precipitated.
3. Place at at -70°C for at least 30 min or -20°C for two hours to overnight.
* 5M NH4OAc, pH 7.4, NaCl and LiCl may be used as alternatives to NaOAc. DNA also may be precipitated by addition of 0.6 volumes of isopropanol.
The key component in DNA precipitation is the centrifugation step. Be sure that centrifugation occurs at least 12,000 x g for 15 min or more.
B. Oligonucleotides
Add one-tenth volume of 3M NaOAc, pH 6.5, and three volumes of cold 95% ethanol. Place at -70°C for at least one hour.
C. RNA
Add one-tenth volume of 1M NaOAc, pH 4.5, and 2.5 volumes of cold 95% ethanol. Precipitate large volumes at -20°C overnight. Small volume samples may be precipitated by placing in powdered dry ice or dry ice-ethanol bath for 5 to 10 min.
D. Isobutanol concentration of DNA
DNA samples may be concentrated by extraction with isobutanol. Add slightly more than one volume of isobutanol, vortex vigorously and centrifuge to separate the phases. Discard the isobutanol (upper) phase, and extract once with water-saturated diethyl ether to remove residual isobutanol. The nucleic acid then may be ethanol precipitated as described above.
