PCR/single 3' mismatch

Garth I. Patterson patterson at frodo.mgh.harvard.edu
Mon Nov 14 09:54:27 EST 1994


In article <3a1ebq$7fv at dns1.NMSU.Edu>, smori at nmsu.edu (Shahram Mori) wrote:

> kenneth garson (cancer research) (kgarson at labsun1.med.uottawa.ca) wrote:
> 
> : 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?

Kwok et al, (1990) Nucleic Acids Research 18:999-1005 is a good reference.
They tried all 16 3' mismatches, and found that the efficiency of the 
amplification depended very much on the nature of the mismatch.  For your case,
only the C:C mismatch will be inhibited, C:A and C:T will amplify as well
as C:G.  For a review, see Erlich and Arnheim, Ann. Rev. Genet 1992 26:479:506.

Garth



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