Nonspecific riboprobe detection of 28S Ribosomal RNA?

[Fred Boyd, Ph.D.]

In article <2i0ngo$l9i at net.usuhs.mil>, denton at usuhs.ucc.usuhs.nnmc.navy.mil
wrote:

> In Article <2hvdb4$e4o at mserv1.dl.ac.uk>
> dmsander at mailhost.tcs.tulane.edu (David M. Sander) writes:
> >
> >Can anyone think of a reason why any particular riboprobe (32-P, 780 bases)
> >might show cross-reactivity with the 28S ribosomal band in a northern
> >blot/hybridization of total rat RNA?
> >
>   I have broad experience in this department.  Unfortunately, it is
> an extremely common and annoying problem particularly with probes with
> GC compositions > 60%.  While there is no other solution for the
> blots other than the one that you have already deduced, (I'm certain
> that you have tried the obvious-increasing the hyb. stringency and
> formamide concentrations) you might consider trying an RNase protection
> assay, if detection of the messaage is all that you require.  I have never 
> had any problem with these  probes and GC content in such an analysis.   
We have recently cloned a bit of DNA with several regions of homology to
28S.  One regions is 51/53 nt identity.  In reading some limited reviews of
28S DNA, the genes are described as "recombigenic," which I extrapolate to
suggest that 28S rRNA may recombine with a variety of other GC rich cDNAs. 
Any comments?
-- 
Fred Boyd                         Voice- 612-624-8150
Lab. Med. Path.                   Fax- 612-616-2444
Cell Biol. Neuro.                 Lab- 612-624-8154
Box 609, UMHC   
University of Minnesota           E-Mail  fredboyd at bmec.micro.umn.edu
Minneapolis, MN   55455