Ethanol Precipitation

Contents

1. Reagents
2. Procedure

1. Reagents

• 3M Sodium Acetate
• 100% ethanol
• 70% ethanol
• TE buffer

2. Procedure

All steps completed on ice.

1. Add 1/10 volume of 3M sodium acetate to each sample.
2. Add 2.5x volume of 100% ethanol to each sample.
3. Incubate @ -20C for at least one hour; overnight is best. Also chill 70% ethanol solution at this time!
4. Centrifuge samples at max speed, <4C for 10 minutes. This must be a refrigerated centrifuge, not just one in a refrigerator.
5. Pipette to remove supernatant, taking care not to touch the sides of the tube. You may not see the pelleted DNA at this time.
6. Add 1mL of ice-cold 70% ethanol to each sample.
7. Centrifuge samples at max speed, <4C for 10 minutes. This must be a refrigerated centrifuge, not just one in a refrigerator.
8. Pipette to remove supernatant, taking care not to touch the sides of the tube. You may not see the pelleted DNA at this time.
9. Dry the DNA on your desktop, or in a speedvac (Ellington lab) on low heat setting for tens of minutes, removing samples as they dry.
10. Add TE buffer to each tube. The more you add, the more dilute it will be; I usually use 20uL.
11. Repeatedly pipette the solution onto the sides of the tube to assure the DNA (stuck to lower part of tube wall) is solubilized again.
12. Check concentration on a nanodrop.