No-boil Chelex 100 proteinase K genomic extraction.

I previously posted a Chelex 100 extraction protocol that while easy, cheap, and moderately effective calls for a boiling step to degrade the proteinase. The degradation step denatures template DNA causing it to become a single stranded product, which is unsuitable for genomic work and in my experience increases the difficulty of PCR.

A protocol published in Molecular Ecology Resources by Casquet et al. (2012) removes the boiling step. Their published protocol allows for high throughout and minimal cost. To their protocol I have added a grinding step which I believe increases DNA yield. I have successfully extracted and used DNA from just 1 leg of a microlepidopteran species.

 

Modified Chelex without boiling from Casquet et al. (2012)

  1. Add 10ul of protinease K (20 mg/ml) to each tube.
  2. Add 150ul of 10% Chelex 100 to each tube.
  3. Grind specimens with melted and sterilized pipet tips
  4. Incubate for 24 hours at 55 °C., swirling occasionally.
  5. Spin down to pellet the Chelex, pull DNA from top.

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