Taq Pol and 3' primer mismatch?

Rob Slebos slebos at amc.uva.nl
Sun Sep 25 17:49:58 EST 1994


In Article <35ni8g$7q8 at louie.udel.edu>
landel at helios (Carlisle Landel) writes:
>
>Will Taq polymerase work if the primer has a 3' mismatch of a single base?
>How many bases of mismatch does it take before the polymerase won't work?
>Does anybody know a reference?
>
>We are trying to PCR a specific member of a highly conserved multi-gene
>family.  I seem to remember hearing that Taq absolutely requires a 3'
>match in the primer to work, but I don't know for sure that it is true.
>
>Thanks for your help!
>
>Carlisle Landel
>landel at helios.medsci.udel.edu

Priming at the 3'-end has been tried to detect point mutations in ras genes,
but does not work very well to distinguish between a perfect match and one
base mismatch. The priming is dependent on nucleotide concentration: at higher
conc. Taq will go over the mismatch. Only when the last two bases don't match
no priming will occur (Cha RS,Zarbl H,Keohavong P,Thilly WG, Mismatch
amplification mutation assay (MAMA): application to the c-H-ras gene. PCR
Methods Appl 1992 Aug;2(1):14-20)

Good luck.

Rob.

Rob Slebos
Academic Medical Center
Dept. Pathology
University of Amsterdam
slebos at amccca.amc.uva.nl





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