PCR: mutations in the primer region?

Warren Gallin wgallin at gpu.srv.ualberta.ca
Thu Sep 3 09:17:39 EST 1998


In Article <35EE6D45.1C02C96 at chclu.chemie.uni-konstanz.de>, "Frank O.
Fackelmayer" <fof1 at chclu.chemie.uni-konstanz.de> wrote:
>Just a thought: Is is possible that Taq makes more errors close to the end of
>a linear DNA than in the middle? 
>What I think of is the following: Usually a polymerase dissociates from the
>template after a misincorporation. Sometimes, the polymerase molecules will
>bind again and extend the strand, giving rise to a "fixed" mismatch. In other
>cases, it will not bind again but leave the strand unextended. Now, if the
>error is "in the middle" of a PCR product, the non-extended strand will not
>amplify any more. If, however, the misincorporation is in the primer region,
>the strand ending with a misincorporated base might very well serve as a
>template for amplification. Over many cycles, you could have more errors in
>the primer region than in the internal sequence. 

Frank,
    The trouble with this idea is that mistakes in the primer region are not
replicated.  That is the part of the PCR product that comes directly from
the primer.  Therefore, any misincorporation in the primer region will not
be propagated into the next generation.

If you find sequence errors in the primer region itself, the problem is
almost certainly an error in the primer synthesis.


Warren Gallin
Department of Biological Sciences
University of Alberta
Edmonton,  Alberta     T6G 2E9
Canada
wgallin at gpu.srv.ualberta.ca



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