T7 transcribed RNA probes

martin LEACH leach at bu.edu
Wed Jan 17 15:27:11 EST 1996


I agree with the last post.

At least run your linearised plasmid out on a gel and see if you got complete
linearization.

Martin


: >From: Torsten Boerchers <borcher at uni-muenster.de>
: >Subject:T7 transcribed RNA probes
: >Date: 14 Jan 1996 13:45:46 GMT
: >Message-ID:<4db1ea$1cpi at majestix.uni-muenster.de>

: >Hello Northern or in situ hybridization specialists
: >
: >Anybody else using T7 RNA polymerase to transcribe
: >RNA from a DNA template (e.g. linearized plasmid with T7-
: >promoter) who also observes multiple bands in a gel or blot?
: >
: >We use this technique for generation of DIG labelled 
: >RNA probes and would like to know whether some 
: >people have explanations for this behaviour or 
: >are aware of published evidence.
: >
: >In some manuals it is emphasized not to allow 34overhangs
: >in the restriction made for linearization. The overhang 
: >could be an initiation point for the T7 RNA polymerase. 
: >Any mechanism for this known?
: >
: >Thank you for any help
: >
: >Torsten
: >------------------------------------------------------------------------/
: >                          Torsten Boerchers                            /
: >    _/  _/_/_/ _/_/_/    Institute of Chemical- and Biochemical Sensor/
: >   _/  _/     _/   _/   Research (Molecular Biology Group)           /
: >  _/  _/     _/_/_/    Mendelstr 7, D-48149 Muenster, Germany       /
: > _/  _/     _/    _/  Fax: +49-251-980 2890 Phone: +49-251-980 2880/
: >_/  _/_/_/ _/_/_/_/  E-Mail: borcher at uni-muenster.de              /
: >-----------------------------------------------------------------/
: >
: >




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