Cell Transformation Group:Protocols/RNA and DNA

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ETHANOL PRECIPITATION OF DNA

Materials:

• 3 M sodium acetate pH 5.2 or 5 M ammonium acetate
• DNA
• 100% ethanol

1. Add 1/10 volume (of DNA sample) of 3 M sodium acetate, pH 5.2, or an equal volume of 5 M ammonium acetate. Mix well.

2. Add 2 to 2.5 volumes of cold 100% ethanol (calculated after salt addition) and mix well.

3. Place on ice or at -20°C for >20 minutes.

4. Spin at maximum speed in a microfuge 10-15 min.

5. Carefully decant supernatant.

6. Carefully add 1 ml 70% ethanol. Invert tube a couple of times to wash pellet.

7. Spin briefly if pellet too loose to tip off 70% ethanol straight away.

8. Tip off ethanol.

9. Spin again and pipette off last bit of 70% ethanol. Air dry tube up-side-down on tissue paper for a couple of minutes. Important not to over dry pellet or will be impossible to dissolve.

10. Resuspend pellet in the appropriate volume of TE or water by pipetting up and down. Can aid DNA resuspension by incubating at 65°C for 10-15min or at 37°C for longer.