Hi all, I'm trying to isolate some gDNA from HEK 293 cells for use in PCR reactions and was wondering if anyone could suggest a method for lysing the cells that I could use without buying in a commercial kit (such as the QIAamp Mini DNA kit)? I've tried lysing them using a 1% Triton X-100 solution but this yielded no gDNA.
If its for PCR, just wash the cells with standard pbs, resuspend the cells in pbs, take about 10^3 cells in 10 ul
add 90ul distilled h20, boil on pcr at 95 for 10 min cool to 4c
and use 1ul of this as ur template for pcr...it should work..you might see a slight background as smear...but as long as ur primers as specific it should be good. Its a lazy protocol but works for just checking.
Hi, thanks for the responses. Chetanchandra, can I just ask what molarity would be best for the Tris and Tris-HCl?
Realised I never replied to you, just to let you know that the protocol worked perfectly thank you =]
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