PCR and Primer Dimer Question

Martin Leach leach at mbcrr.harvard.edu
Tue Oct 5 12:31:11 EST 1993


In article <1993Oct4.180524.397 at ucsvax>, Rodney Tweten
<Rod-Tweten at uokhsc.edu> wrote:

> Hello,
> I was wondering if someone could tell me how much of a problem primer
> dimers are in PCR.  I am performing PCR mutagenesis on a gene that is AT
> rich (about 70%) and it is almost impossible to get away from primer
> dimers.  For instance, the worst case is one where I have 8 bases out of
> 11 showing hybridization in a 22-mer.  The bases are all A-T pairs. 
> Other examples would be 6 out of 7, again all A-T pairs.  I realize that
> primer dimers can kill a PCR reaction but can a hot start eliminate this
> problem?  Any help appreciated.


Dear Rodders,

They can be a real pain in the ass.

Look at the 3' ends of both primers and make sure there is nothing
complementary at the far 3' end of both primers. There is a program called
Amplify that can simulate the PCR reaction and this is obtained by ftp to
ftp.bio.indiana.edu
Have played with the program and it does pretty funky stuff with your PCR
reaction.
You have to play around with text editing to insert your primers, seq etc..
but it works reasonbly well.
It also predicts primer dimers for you!!!!

If you can't find it drop us a line

Martin Leach



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