> To: biochrom at net.bio.net> From: frog <pennie at access2.digex.net>
> Subject: Re: pcr taq pfuu
> Date: Thu, 2 Feb 1995 13:03:22 -0500
>>> On Thu, 2 Feb 1995, Philip L. Carl wrote:
>> > Presumably it does get back on and proceed some considerable fraction of
> > the time-that's why Taq PCR is error prone. The key here is the ratio of
> > exo activity to polymerase activity, and while I agree the ratio of 100:1
> > is a bit surprising, it's not so surprising that I find it a mystery.
> > Phil Carl
>>> I agree but there are other factors which could come into play;
>> 1) How far does taq continue after introducing an error. Does it fall off
> right away or not? If it can proceed further than pfu's ability to
> proofread back then a significant number of errors may go undetected.
>> 2) What is the relative affinity of taq vs pfu for a missmatched
> template/primer. In other words if taq is poorer (say 100 times less
> affinity than pfu) at settling onto a mismatched elongating template
> you can imagine it falling on and off without getting anywhere. Then
> along comes the 1 in 100 molecule of pfu with no such distaste for a
> mismatched base pair and off it goes.
>> If anyone can elaborate on any of these points I'd be happy to know
> (eg relative Km values for correct and missmatched templates, limit of
> "back" proofreading of pfu, processivity of taq after error).
You might check out a paper by Huang, Arnheim and Goodman in Nucleic
Acids Research 20(17):4567-4573. `Extension of base mispairs by Taq
DNA polymerase: implications for single nucleotide discrimination in
They don't compare Taq and Pfu but they do sort out some of your
association/dissociation questions for Taq. Maybe that will help,
||Doug Rhoads || Dept. of Biological Sciences||
||drhoads at mercury.uark.edu || 601 Science Engineering ||
||drhoads at uafsysb.uark.edu || University of Arkansas ||
||501-575-3251 || Fayetteville, AR 72701 ||
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