Kitto Lab, The University of Texas at Austin http://research.cm.utexas.edu/bkitto/Kittolabpage/Protocols/Microbiology/ethanolPpt.html

This procedure allows the concentration of DNA samples from dilute solution and the removal of unwanted salts from DNA samples.

Materials

•3M Sodium Acetate buffer, pH 5.2 (store at 4 ℃)

•Cold 100% Ethanol (-20℃)

•Cold 70% Ethanol in sterile dH₂O (-20℃)

•DNA sample

•4 ℃ Microcentrifuge (normal microcentrifuge in cold room works fine). All centrifugations should be on "soft" (no brake) setting.

Procedure

1.Transfer DNA to a container where it fills one fourth the total volume (a 500 μl tube should have no more than 125 μl of DNA solution, for example).

2.Add one tenth volume of Sodium Acetate buffer to equalize ion concentrations.

3.Add at least two volumes of cold 100% ethanol; let stand in -20℃ freezer for at least one hour .

4.Centrifuge sample for 15 minutes at highest speed in a 4℃ microcentrifuge .

5.Remove as much supernatant as possible with a 1 mL micropipet; recentrifuge, then remove the rest with a 200 μl pipet.

6.Add 200 μl of cold 70% ethanol; centrifuge for 5 minutes in a 4 ℃ centrifuge.

7.Remove supernatant with a 200 μl pipet; evaporate remaining ethanol in a 37 ℃ water bath.

8.Resuspend pellet in desired volume of water or TE buffer.