Ethanol Precipitation

Materials and Reagents

For 20 μL reaction (ligation, recombination, etc.) add:

  • 1 μL of 20 μg/μL glycogen

  • 2 μl of 3 M Sodium acetate (pH=8.5)

  • 50 μl of 100% Ethanol

Procedure

  1. Precool chiller to 4 Celsius.

  2. Put reaction in -80 Celsius freezer for 15 minutes. Longer durations appear to increase yield.

  3. Spin at 14,000 RPM for 30 min.

  4. Wash with 200 μL of 75% Ethanol.

  5. Spin for 15 minutes at 14,000 G.

  6. Wash with 200 μL of 75% Ethanol again.

  7. Spin again for 15 minutes at 14,000 G.

  8. Very gently remove the ethanol, DO NOT disturb the pellet.

  9. Resuspend in desired volume (usually 2.5 μL) of TE (Tris-EDTA).

Notes and Cautions

  • I find that the round-bottomed Eppendorf tubes are the easiest to work with, since they enable facile visualization and air-drying of the DNA precipitate.