filter sterilized vs. autoclaved water for PCR

Dave Knaebel knaebel at draco.clarkson.edu
Thu Apr 2 16:08:25 EST 1998


My lab group has run into some problem recently with getting PCRs to
work in the lab, which had always worked in the past. We tracked down
the problem to the water used in the dilution of the reactions. In the
past, we have successfully used autoclaved 18 megaohm water for this,
but now, this results in complete inhibition of the PCR. Filter
sterilizing the same water results in perfect results.

Does anyone know the mechanism for why this inhibition occurs? Also,
does anyone have any ideas as to changes that can occur in an autoclave
that would result in it leading to inhibitor formation in the water?
Alternatively, some particulates could be being produced (or bled out
of) our water system that are removed during filtration but left in
during autoclaving.

Thanks for any information or ideas.

Dave Knaebel
Biology Department - 5805
Clarkson University
Potsdam NY 13699-5805
(315) 268-2391
(315) 268-6610
http://www.clarkson.edu/~knaebel





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