possible contamination of Taq-polymerase with plasmid sequences

Paul N Hengen pnh at ncifcrf.gov
Tue Mar 25 12:25:03 EST 1997


Georg Kroeger (a03m at biologie.uni-bremen.de) wrote:

> Of course I checked everything, in all combinations. Finally I lend
> Taq-polymerase from a neighbour lab (Pharmacia product), and -
> everything was fine. Using my buffers, templates, dNTP's I got
> expected  results with the Pharmacia Taq, but all positive using Gibco
> Taq.

> I phoned Gibco and they send me an aliquot of another Lot (this time
> recombinat Enzyme) to test. This lot was OK!. I ordered more, and -
> everything went wrong. 
> Once again we searched for a homemade contamination and again the
> Enzyme (not the very lot I tested before) was the bad guy. Now we are
> going to buy our Taq from Pharmacia and hope that one is clean.

There are no guarantees that every lot of Taq from this company will
be free from all recombinant plasmid DNA either. One thing to do
would be to buy several lots and do the negative control for every
tube you get, then choose the best lot for your experiment. Maybe also
try MCS primers on the positive ones and see if you get consistent bands.

> By the way, Gibco said they never had any problems with their Taq.

Of course not! They never test everything for extra DNA added ;-)

> Actually I do not think I am a genius who had this fatal primer pair
> in mind for the first time in human history :).
> So please, is there anyone in the Lab-world who had strange results
> with  beta-Lactamase primers? Any comments will be wellcome!!

Yep, it's happened plenty of times before...

@article{Hengen1994Octtibs,
author = "P. N. Hengen",
title = "Methods and reagents - Better competent cells and
{DNA} polymerase contaminants",
journal = "Trends in Biochemical Sciences",
volume = "19",
number = "10",
pages = "426-427",
month = "oct",
year = "1994"}

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