Use of Salt Gradient in Sequencing Gels?
finney at Frodo.MGH.Harvard.EDU
Tue Sep 14 20:13:38 EST 1993
In article <2756q2$2jd at charm.magnus.acs.ohio-state.edu>, rrumpf at magnus.acs.ohio-state.edu (Robert Rumpf) writes:
>I'd be interested in hearing protocols and experiences from those who have used
>salt gradients to improve sequencing gel results. We have seen a protocol in
>USB's editorial comments 19:2 page 64 that sounded intriguing, but wanted
>second and third opinions. Any help would be appreciated.
The protocol in common use here at Mass General Hospital was originated
by Sheen and Seed (Biotechniques 6:942-944, 1988) and is also described
in Current Protocols in Molecular Biology p. 7.6.8.
The basic idea is to add NaAc to the bottom buffer to a final conc.
of 1M. The effect is to prevent bands that would otherwise run off
the end from going for a swim in the bottom tank. In a typical
run probably 20-50 extra bases are retained. Sometimes this enough
to save you the trouble of doing double loadings.
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