cloning of PCR products - errors?

Michael Witty mw132 at mole.bio.cam.ac.uk
Wed Jun 27 08:32:22 EST 2001


On Wed, 27 Jun 2001, Sergio wrote:

>
>
> "Frank O. Fackelmayer" wrote:
>
> > Kristo K M Kulju wrote:
> > >
> > > "Frank O. Fackelmayer" <Frank at Fackelmayer.de> writes:
> > >
> > > > "Richard P. Grant" wrote:
> > > > >
> > > > > > errors but what about cloning procedure? Is is considered to be
> > > > > > practically "error free"?
> > > > > Yes.  That's why it's called 'cloning'.
> > > > agreed, it is CONSIDERED to be error free. But of course it is not. Bugs
> > >
> > > Thanks for the help! Does anybody know references about this? Something I
> > > could cite if I want to say that Taq-errors are more probable reason of
> > > point-mutation artefacts than cloning.
> > >
> >
> > Using published values for the error rates of Taq polymerase and E.coli
> > polymerase I you could also give a rough impression of the probability
> > that a particular error is due to PCR. Taking the values from the NEB
> > catalog (page 75), the error rate for Taq it 285x10-6, that of PolI is
> > 9x10-6. That is a ratio of around 30.
>
> Only 265x10-6 ??... damn... i must be doing something wrong. We use to get
> around 1x10-3 mutations when using Taq... almost five times the supposed error
> rate.
>
> Sergio

In one experiment I got about 1 error in 300bp BUT in another zero errors
in over 2000bp (both times using Taq).  Mike.




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