colony pcr question

Don Walthers gene_jockey26 at
Mon Jan 10 17:47:25 EST 2000

I routinely perform PCR on colonies of M. xanthus (67% G+C). Take a large
colony and resuspend in 10-20 ul H2O. Test 1-10 ul of template. I usually do
95-5' (95-1', 50-2' and 72-X') X30, with and without 5% DMSO. The low and
lengthy annealing temp has been a critical factor in getting these GC-rich
PCRs to work. Also the difference between 94 and 95 in the denaturing step
is important. Make sure your Taq doesn't mind 95. We use standard BRL Taq.

Don Walthers
B.S.Microbiology U of Illinois at UC, 1996
Ph.D. Microbiology U of Idaho, 2001?
gene_jockey26 at

More information about the Methods mailing list