pcr taq pfuu

frog pennie at access2.digex.net
Thu Feb 2 13:03:22 EST 1995



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). 

Later.........

Frog.



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