Overhang A of PCR

Michael J. Prigge prigge at darkwing.uoregon.edu
Wed Apr 30 20:44:39 EST 1997


In article <GEjpQAAG60ZzEw9m at genesys.demon.co.uk>, "Dr. Duncan Clark"
<duncan at genesys.demon.co.uk> wrote:

> In article <H+7JeKAn5eZzEw+U at genesys.demon.co.uk>, "Dr. Duncan Clark"
> <duncan at genesys.demon.co.uk> writes
> >They all have the
> >associated terminal transferase activity and add preferentially a single
> >A (*) overhang. 
> 
> 
> The asterisk was there for a good reason and I forgot about it.
> 
> There was a paper publsihed in Biotechniques in Vol 20 No. 6, pp1004-10
> that investigated primer design and addition of 'A's. They found that
> Taq prefers to add a terminal base adjacent to a C  ie a 5'G on your
> primers. Having a 5' T on the primers pretty much stops the terminal
> base addition.
> 
Does it follow that I should be making my T-tailed vector using HincII
(GTC/GAC, in pUC at least) rather than EcoRV (GAT/ATC)? or does this 
not hold as well for adding T's?
Michael Prigge                          Phone: (541) 346-4256
Institute of Molecular Biology            Fax: (541) 346-5891
University of Oregon /\\ /\\ /\\ /\\ /\\ /\\ /\\ /\\ /\\ /\\ 
Eugene, OR 97403   \\/ \\/ \\/ \\/ \\/ \\/ \\/ \\/ \\/ \\/ \\



More information about the Methods mailing list