Help!PCR problems.

Klaus Salger salger at wap18.zi.biologie.uni-muenchen.de
Sat Feb 18 14:08:56 EST 1995


sl1nn at cc.usu.edu wrote:
: I am having problems with PCR amplification of a small fragment of cloned
: DNA.  I
: I have designed my own primers, using Oligo 4.1, and according to the
: program I
: am using the right temperatures for the primers. I have varied tha
: conditions
: several times with the same results.  The negative control (without
: template) of my two primers always shows a 250 bp band.  Both primers
: are
: 22-mers and the dimers would not be that large. In additon, when I have
: the
: template present I am only seeing the same band. I have checked the
: primers for
: contamination using HPLC, and they check out at the right size and with
: only
: one peak.  Does anybody have any idea why I am getting this result?  I'm
: going
: crazy trying to fix the problem.   Thanks in advance.

Band without template, no contamination in the primers - maybe something
else is contaminated. As far as I remember there was some discussion
about DNA contaminations in Taq preperations and how to get rid of them.
You could search the old articles on "gopher://net.bio.net".
Hope this helps
  Klaus

--
Klaus Salger                phone : ++49 (0)89 5902 -502
Zoologisches Institut       FAX   :                 -450
AG MacWilliams              e-mail: salger at zi.biologie.uni-muenchen.de
Luisenstr. 14
80333 Muenchen
Germany



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