PCR probes by incorporation of 32P ?

Robert Horton horton at biosci.cbs.umn.edu
Mon Mar 4 14:45:58 EST 1996


Geoffrey Neale (geoffrey.neale at STJUDE.ORG) wrote:

: Hello everyone.

: I am looking for a method for making 32-P labeled probes during PCR. The
: information that I specifically need is the concentration of cold 
: nucleotide in the reaction so that you generate sufficient full length 
: probes, but still retain high specificity of the isotope label.

: Other considerations are the amount of starting template, the 
: concentration of primers, and the number of cycles.

: Thanks for your input.

: Geoff Neale

: PS Could you please email me your advice since our news reader is not yet 
: operational on our system. E-mail is my only access at this time. Thanks.

(Yeah, I'll e-mail it, too. Posting here for the general audience...)

See:

Schowalter DB.  Sommer SS. The generation of radiolabeled DNA and 
RNA probes with polymerase chain reaction.  Analytical Biochemistry.  
177(1):90-4, 1989 Feb 15.

You can achieve higher specific activities with lower total [dNTP],
but you need to increase the extension time.

---
Robert M. Horton
http://134.84.47.3                                       Have a :) day

"Scotty, try flushing the radioactive waste into the ventilation system!"



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