Taq error problems

Tracy Aquilla aquilla at salus.med.uvm.edu
Tue Jan 17 14:11:07 EST 1995


In Article <D2Jspv.H2u at info.swan.ac.uk>, G Jenkins wrote:
>I'm using a PCR technique to detect mutations in genomic DNA
>in order to quantitate the mutations I need to detect a very
>small number of mutations above a Taq error background. I
>therefore need to reduce the Taq error rate as much as possible. I 
>know about the addition of a proofreading polymerase, and am trying 
>that at present. Can anyone help with any other suggestions, e.g.
>cycle number (I'm using 24), DNA concentration ( I'm using 10 microg's),
>primer concentration (20 pM)?
>Any help would be greatly appreciated
>Gareth Jenkins

1. keep MgCl2 conc. as low as possible while still giving decent yield
2. keep number of cycles as low as possible too
3. keep dNTP conc. as low as possible, and no higher than MgCl2 conc.
Good luck

Tracy Aquilla, Ph.D.
Molecular Physiology and Biophysics
University of Vermont
aquilla at salus.med.uvm.edu



More information about the Methods mailing list