PCR mistakes

Richard P. Grant see_sig at cmtech.co.delete.uk
Wed Jun 3 05:02:34 EST 1998


Hi Clemens

I found that I could get good error rates (i.e. no mistakes) with Taq by
lowering the concentration of dNTPs, from 200 uM to 50 uM.

Using this method, 1.9 kb frags amplified with no worries.  You may end up
with less product, admittedly . . .

R

In article <6l35jo$6lk at sun0.urz.uni-heidelberg.de>,
un691cs at genius.embnet.dkfz-heidelberg.de (Clemens Suter-Crazzolara, PhD)
wrote:

: Hello,
: 
: I am trying to clone a PCR product of 750bp, but it appears to
: contain about 4 PCR mistakes. The polymerase I use is Ultma from 
: Perkin Elmer. 
: As the RNA is of pretty low abundance, I have to amplify for 32 
: cycles, still this wasn't much of a problem in the past.
: 
: Do you have any idea how to circumvent this high failure rate of
: the PCR ? (I re-call lowering the MgCL2 concentration, currently
: at 2.5 mM).
: 
: Thanks, clemens

-- 
Richard P. Grant MA DPhil  | rgrant at  cmtech.co.uk
Senior R&D Scientist       | work: http://www.cmtech.co.uk/
Cambridge Molecular        | home: http://www.avnet.co.uk/adastra
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