High G/C PCR

Greg Nattrass gnattras at waite.adelaide.edu.au
Wed Feb 5 07:54:45 EST 1997


I've been battling the same problem for about the last 6 months.
However, the GC rich sequence I must PCR through is preventing me from
obtaining a 5' RACE product. I have a segment of cloned DNA which
contains 80% GC (over 200bp) and up to 90% in some 50 bp segments. The
only way I have been able to amplify across this region with Taq and
Elongase (Life Tech), has required NaOH denaturation of the cloned DNA,
ethanol precipitation then PCR. I have to use longer denaturation times
as well ie 1 min 30 sec. If you require more help or an article
explaining this procedure feel free to email me.

Greg Nattrass



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