Beta-Actin good RT-PCR Control?

David Huen smh1008 at cus.cam.ac.uk
Fri Dec 30 16:00:18 EST 1994


In article <3e1ioo$1nn at netnews.upenn.edu>, smoore at mail1.sas.upenn.edu (Sean
David Moore) wrote:

> Dear BioNetters....
> 	Here's a situation for you.  I have been isolating human alveolar 
> macrophages from sick patients.  I treat them with different experimental 
> proteins and later look for message of cytokines.  I have lately had 
> trouble finding beta-Actin as a positive RT-PCR control..I think there 
> may be two problems:
> 
This might be a bit of a dumb question but it has puzzled me for a long
time. How do you use beta-actin as an RT-PCR control when genomic DNA has
beta-actin pseudogenes (it does, doesn't it?)? Surely under these
circumstances, irrespective of the integrity of your mRNA and success of
your RT reaction, there should be enough genomic DNA carryover to at least
occasionally generate a false positive in RT-PCR.

David Huen aka Puzzled, Dept. of Pathology, Univ. of Cambridge.



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