PCR/single 3' mismatch

kenneth garson cancer research kgarson at labsun1.med.uottawa.ca
Wed Nov 9 16:33:05 EST 1994


I have a PCR template which is degenerate at at least one position. I
would like to amplify specifically those templates which contain a G at
this position. If I design a PCR primer whose 3' base matches a "G" at
this degenerate site, could I be assured of only amplifying templates
which contain a "G" at this position, or will annealed primers with a
single 3' mismatch have a probability of amplifying? Cloning is not really
a favourable option since the amount of available template is low, so PCR
is necessary, and the percentage of template harbouring a G at this site
may be low so screening clones of PCR products using external primers
would be  a lot more work.  Any ideas? Any references?

Thank you for any input.


Ken Garson
Ottawa Centre for Cancer Research
University of Ottawa,
Ottawa, Ontario
Canada
K1H 8M5
kgarson at labsun1.med.uottawa.ca



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