Why do probes stick to RNA markers?
oberstra at hrz.uni-kassel.de
Fri Sep 29 04:34:20 EST 2000
> > Many, but not all, of our 32-P labelled probes stick to the marker lane
> > on our northern blots. This happens with Sigma and Gibco BRL RNA ladders
> > and the bands the probes hyb to don't correspond always to the marker
> > bands.
> > Does anyone else get this, and can anyone give an explanation as to
> > whats happening?
> > If you can help us out with this query e-mail me at
> > jonathan.howarth at bbsrc.ac.uk
> > Thanks for any ideas
> > Jonathan
Do you hybridize with in vitro transcripts or with oligo-labelled DNA?
In general the markers are produced by companies using cloned inserts.
Because nearly all trancription vectors share common sequences around
the MCS the part of your probe containing MCS sequences fits to the
marker. There are several ways to avoid this signals (use PCR product as
template, use RNA as probe, cut exactly beside your insert etc.), but we
love them because they make the length estimation easier.
Juergen Oberstrass E-Mail: oberstra at hrz.uni-kassel.de
Kassel University Genetics Department
Heinrich Plett Str. 40
D-34132 Kassel / Germany
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