Ethanol Precipitation of Nucleic Acids

Materials

  • Cold 100% ethanol
  • Cold 70% ethanol
  • 1 M sodium acetate
  • Centrifuge cooled to 4°C
  • Heat block warmed to 37°C

Protocol

  1. Transfer nucleic acid to a container where it occupies less than one quarter of the total volume (e.g., a 1.5 mL tube should have no more than 375 μL of nucleic acid solution).

  2. Add one tenth of the nucleic acid volume of sodium acetate buffer to equalize the ion concentrations.

  3. Add 2-3 volumes of cold 100% ethanol and place the sample in a -20°C freezer for at least one hour.

  4. Centrifuge sample for 15 minutes at 12000 x g at 4°C.

  5. Remove as much of the supernatant as possible. Be careful not to disturb the pellet. If you get all the liquid out then skip step 6.

  6. If not then re-centrifuge briefly, then remove the rest with a 200μL pipette.

  7. Add 250 μL of cold 70% ethanol.

  8. Centrifuge for 5 minutes in a 4°C centrifuge at maximum speed.

  9. Remove supernatant with a 200 μL pipette; evaporate remaining ethanol in a 37°C heat block. Do not do this for too long because overdrying the pellet can make it difficult to resuspend.

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