High G-C PCR...help.

Dr. Duncan Clark duncan at genesys.demon.co.uk
Thu Apr 18 03:45:51 EST 1996


In article <Dq0MAE.367 at murdoch.acc.Virginia.EDU>, Steven James Carlisle
<sjc4h at faraday.clas.Virginia.EDU> writes
>I need advice on PCR...I've tried many times to amplify
>a 1.2 Kb insert with multiple primers, different enzymes,
>different templates, different TM's, and lots of bad
>language...nothing.
>It has a very high (about 75%) G-C content...is this a problem?
>Anybody else had this experience?
>

Avoid PCR buffers with KCl as it stabilizes secondary structure (a
recent NAR single sheet at back paper). Try DMSO ie 2% or 5% final,
extra glycerol, 1% Tween20 final or other non-ionic detergents. 

Duncan
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