Biology Protocols

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Phenol/Chloroform Extraction of DNA

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1. Add an equal volume of buffer-saturated phenol:chloroform (1:1) to the DNA solution to be extracted.

2. Mix well by vortexing for 5-10 seconds.

3. Centrifuge at >10,000rpm for 5 minutes.

4. Carefully remove the top aqueous layer to a fresh tube, avoiding the interface between the layers (if necessary repeat steps 1-4 to maximise yield).

5. To remove phenol traces from pooled aqueous layer, add an equal volume of chloroform to the aqueous layer.

6. Centrifuge at >10,000rpm for 5 minutes.

7. Carefully remove the top aqueous layer to a fresh tube.

8. Concentrate the aqueous layer using an ethanol precipitation protocol.


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