Additives and PCR Fidelity

vilimf01 at mcrcr6.med.nyu.edu vilimf01 at mcrcr6.med.nyu.edu
Thu Jun 13 19:29:37 EST 1996


Hi Netters,
	I am posting this because of recent discussions on the use of
DMSO, formamide, and other additives in PCR.  I have used these with
varying degrees of success with taq and vent.  The problem is that the
subsequent sequencing of these products revealed an unacceptable error
rate ( approx 4-6/kb even with vent ).  This is a bit disconcerting. 
I am wondering if these additives cause decreases in fidelity or if I
am picking up the mutations in RT or gel purification/UV
visualization.  As far as I know, no one has looked at the effects of
these additives on fidelity.  Can anyone comment on their experiences
with these (and/or other) additives with regards to the error rates
they see.
	The other thing that bothers me is how can a bacterium
(thermus aquaticus) survive with such an error prone DNA polymerase. 
Maybe the polymerase isn't so error prone, but conditions for
improving its fidelity have never been optimized.  Any comments?

				Regards			SVEN



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