PCR screening: False positives

Dr. W. Greenhale wgreenha at hgmp.mrc.ac.uk
Tue Sep 6 11:38:57 EST 1994


Hi, a colleague of mine recently came across what I 
considered to be a fairly worrying phenomenon.  
We routinely PCR screen E.coli colonies in our lab 
and sometimes find PCR positives which are 
\expression negative (not very surprising).  However, 
it seemed that this was more common after 
electroporation.  Acting on a hunch my colleague 
therefore tried PCR-ing (I take it the verb to PCR 
is now official English grammar at least on the bionet) 
up random areas of colony free regions on her plates 
and came up with a remarkably high level of PCR 
positives.  Obviously the reason is that traces of the 
transforming DNA were present on the plate.  This can 
be overcome by carefully picking the centre of colonies 
(not always easy especially in a big screen).  What I 
was wondering is if this problem is common knowledge 
and if not shouldn't people be warned.  I for one had total 
faith in the PCR screening technique and have almost 
certainly wasted quite extensive periods expressing 
negatives.

Bill Greenhalf
MRIC Biotech.
NEWI - Deeside,
Connah's Quay
Clwyd, UK

E.mail Greenhalfw at newi.ac.uk
or as above



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