preferential amplification by proofreading Taq ???
ed at bio02.bio.uottawa.ca
Sat Mar 22 14:50:44 EST 1997
thanks to all who've replied...this is crazy but here's what's happened
I set up some duplicate reactions using both Taq and TAKARA Taq, and this
PCR seems a little on the iffy side. Whereas before only Taq worked, this
time one out of two Taq reactions amplified the sequence of interest, and
surprisingly (perhaps because of some cosmic realignment of nebulae far,
far away), both TAKARA reactions amplified the sequence of interest. The
stuff is getting cloned as we speak...
ah, nothing like experiments that you can't replicate...
On Sat, 22 Mar 1997, DHolst6016 wrote:
> It is strange that you have problems amplifying the same size product with
> Taq and proofreading Taq. One would suspect nonspecific priming. One
> thought my be that since Taq polymerase has such a broad activity range,
> 30-85C, nonspecific priming may be an answer. Possible solutions may be
> hot start PCR or use af a proofreading enzyme such as Pfu which has a much
> tighter range of activity, hence less nonspecific product.
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