Northern Problem

Steven Coon sdcoon at umich.edu
Sat Mar 26 23:22:36 EST 1994


In article <199403252125.NAA05726 at net.bio.net>,
KGEISS%RAVEN at EAGLE.AAMRL.WPAFB.AF.MIL wrote:

> We are currently having problems with our northerns.  It seems that we get a
> lot of nonspecific binding of our probes to both ribosomal RNA bands.  We are
> using cDNA probes for cytochrome P450 2E1.  Any insight into this problem is
> appreciated.  Post or e-mail responses.
> Thanks,
> Geiss...[:)


After hybridization you are not washing them either long enough, hot
enough, or the SSC concentration is not low enough. It is also possible
that the unincorporated radioactive NTPs are not being removed well enough.
What kind of probes are you using? riboprobes tend to do this more often
that cDNA probes (don't know why). Try raising the specific activity of
your probes and washing your blots more stingently.  



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