Hot start PCR
Dr. Duncan Clark
Duncan at nospam.demon.co.uk
Thu Aug 19 10:41:19 EST 1999
In article <37BBF881.43DF51C4 at pitt.edu>, Rich Dudley <rdudley+ at pitt.edu>
>For Amplitaq gold (PE) and Platinum Taq (Life Technologies), they use monoclonal
>antibodies directed against the active site.
Not for AmpliTaq Gold only Platinum Taq.
AmpliTaq Gold is chemically modified and has a patent granted for the
side chain modification agent they use along with claims for any other
agents of the same chemical structure. The modification is pH sensitive
as in reversible at low pH i.e. as you take your Tris PCR reaction
buffer from RT to 94C it's pH drops sufficiently low enough to
reactivate the pol. Very neat and darned difficult to find something
that has a similar effect but is outside the patent claims.
Then again it has proved quite difficult in house to force a PCR to fail
such that hot start is needed. Spiking v.low copies of a target into
human genomic has not reliably produced results similar to those
published in the original antibody or oligo inhibitor papers.
The problem with being on the cutting edge is that you occasionally get
sliced from time to time....
More information about the Methods