Quick PCR ques.

[Alex Dobrovic]

From: dcarpio at mail1.sas.upenn.edu (David Carpio)

>I'm having a few problems with my PCR protocol, and I've heard that
>having *too many* dNTPs can inhibit the reaction.  I was wondering what the
>reason for that would be.
>
>Any thoughts?
>
>Dave

dNTPs bind Magnesium stoiochiometrically. Taq polymerase needs free Mg++ as
a cofactor. Typically with 0.m8M dNTPs and 1.5 mM, there is 0.7mM free
Mg++. (actually a bit less as the DNA binds some too). If you were to
double the dNTPs, there would be no free Mg++.


Alexander Dobrovic, Ph. D.
Chief Scientist
Department of Haematology-Oncology
The Queen Elizabeth Hospital, Woodville, SA 5011, Australia

Affiliate Senior Lecturer
Department of Medicine, University of Adelaide

Tel 61-8-8222 6884 Fax 61-8-8222 6046