Factors affecting the effciency of end-labelling by DNA kinase

[Hiranya Roychowdhury]

At 04:44 PM 10/21/96 GMT, Dominic Voon wrote:
>Hi all,
>	I am going to start labelling some PCR products using DNA kinase
>(with alpha-32P dCTP) and it is quite important that I get high
>effeciency. I am just wondering if anyone could just give me a quick
>run-down on the critical factors that contribute to the effeciency of this
>experiment ? such as DNA concentration, dCTP concentration etc. Any
>pointer/comment/suggestion/advice will be much appreciated. Please email
>me directly at dvoon at cyllene.uwa.edu.au.
>
>many thanks in advance,
>dominic


I am not sure you know what you are doing! Kinase will transfer the cold
gamma-phosphate group from the dCTP. You sould use gamma-32P-ATP; that's
what I  ( and I believe, most) do. There is really nothing else that could
go wrong if you follow a standard rxn protocol.
Dr. Hiranya Sankar Roychowdhury
Plant Genetic Engineering Lab.
New Mexico State University
Las Cruces, NM 88003
Ph. (505) 646-5785
hroychow at nmsu.edu