DNA Contamination in PCRs

Marko Rehn rehn at phoenix.oulu.fi
Sun Jun 27 07:59:52 EST 1993


Hi !

I encountered the same problem about two years ago. Two useful articles are
in BioTechniques (1991) Vol. 10: No.1, 26-30; No.4, 442-445. I have used 
the UV-irradiation method succesfully. All you have to remember is that not
to have adjacent Ts in your primer. Another problem is buffers containing
Triton (Promega). When we changed to Promega Taq I noticed that when I
irradiated the buffer along with others (water, primers, dNTPs) there was
not any product. When the buffer was left without irradiation the product
was synthetized. The solution may be that Triton (aromatic => absorbs UV
strongly) cross-links to primers or dNTPs ??

When the contaminating fragment is very short (< 300bp) the UV-method might be
not enough (only few neighboring Ts). In that case the restriction digestion
method (the first reference above) might be useful. In my hands the digestion
reaction and the PCR have worked in the same buffer (fg. NEB 3).

Hopefully this helps.

Marko Rehn
Department of Medical Biochemistry
University of Oulu
Finland

rehn at lbiok-1.oulu.fi
rehn at phoenix.oulu.fi
mrehn at csc.fi



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