How to avoid PCR errors?
roth at husc7.harvard.edu
Thu Feb 10 19:22:48 EST 1994
In article <oliver.4.2D5A3C2F at molgen.biologie.uni-marburg.de> oliver at molgen.biologie.uni-marburg.de writes:
>I want to amplify 2 kb genomic DNA and then to subclone the DNA (actually I
>have to amplify it two times, because I perform a site directed mutagensis).
>It is essential, that the sequence of the subcloned DNA is the SAME as the
>original sequence. Until now I did not manage to get a clone without PCR
>Has anyone experience with such a problem and knows how to circumvent this?
>I think there are some polymerases wich have a proof reading function. Has
>anyone ever used such an enzyme and can tell me wich is the best to use for
>Any help is highly appreciated,
> Novell-Server Molekulargenetik Biologie Uni-Marburg Deutschland
Check out Pfu.... Lundberg et al., Gene 108:1-6 (1991)
Pfu (from Pyrococcus furiosus-- 'rushing fireball') is a proofreading
polymerase w/ 1 or 2x10^-6 error rate.
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