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PCR to measure load

Todd Miller - Pharmacology tmiller at newssun.med.miami.edu
Fri Mar 31 14:30:41 EST 1995


	I'm not a virologist but I have had some experience
with PCR.  I had a question that I hope someone can help me
with.  It seems that PCR has now been found in several systems
to consistently measure ~2-4 logs more "virus" than the
traditional infectivity assays.  Often these assays simply
involve doing the PCR on a "viral pellet".  In the case of
HIV, which I'm most interested in, I think the primers are
targetting the gag sequence.
	If I remember my limited training in virology correctly,
I thought that when viral proteins were made, in many cases
a processed RNA transcript is the template.  By using the
gag primers that everyone uses to measure viral load of HIV,
can one really distinguish whether the original template was
actually part of an intact virus, or could processed viral
message that happened to contaminate the "viral pellet" also
be a potential template?  I was wondering if primers could
be designed that would only amplify unprocessed (ie, destined
to be or already part of a viral particle) RNA.  These would
probably produce a product that spanned 2 or more viral
message regions.
	Thanks for any help.

Todd Miller
tmiller at newssun.med.miami.edu




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