Help! RNA Protection Assay

Anton Tutter atutter at aim.salk.edu
Fri Jun 21 00:50:28 EST 1996


Hello Netters:

I need to quantitate transcripts of varying lengths off of a given promoter (HIV-1 LTR) in 
vitro.  Several techniques have been employed by other groups with varying degrees of 
success:  S1 Nuclease protection, RNase protection, g-less cassettes.

Specifically, I need to be able to quantitatively detect transcripts from 0 to >700nt in length.  
I am considering making full-length anti-sense RNA probes by asymetric PCR of my 
promoter construct.  Is 700nt too long for a ss anti-sense probe?  i.e., will that introduce 
complications in hybridizing to the transcripts?  

Has anyone reading this done something similar?  I would appreciate all ideas and input.  
Please e-mail me, as my news server is often down.

Thanks in advance,

Anton
atutter at aim.salk.edu



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