Team:Cambridge/Protocols/Extraction of genomic DNA from squid

Extraction of genomic DNA from squid

A method to extract genomic DNA from squid tissue Large sample number, very quick and dirty, adequate for PCR

protocol adapted from http://zfin.org/zf_info/zfbook/chapt9/9.3.html

Theory

How it works

Practice

This protocol is most suitable for samples consisting of 1-20, diploid, 2-3 day old embryos. Embryos need not be removed from their chorions. DNA prepared with this procedure is only good for PCR analysis, but is unsuitable for digests and Southern blots. Because PCR does not require high-molecular-weight DNA, samples can be vortexed and frozen.

1. Transfer embryo(s) into microfuge tube and remove excess liquid with drawn-out Pasteur pipette.

2. Add extraction buffer, 50 µl or 10 µl per embryo whichever is larger, and incubate at 50-56°C for 2-3 hrs (longer is ok). Vortex occasionally.

PCR Extraction buffer

10 mM Tris pH 8 2 mM EDTA 0.2% Triton X-100 200 µg/ml Proteinase K

3. Boil samples in waterbath for 5-10 min to inactivate Proteinase K.

4. Spin in microfuge for 1 min, store at - 20°C.

5. Use 10-15 µl to set up PCR reaction in a total volume of 50 µl, proceed with your favorite PCR protocol.

Safety

The safety implication of the procedure.