Some questions about Polyacrylamide gels

Fabius FabioDB#deleteme# at katamail.com
Sun Mar 7 09:58:11 EST 2004


Hello.
I use to do little vertical (10X10 cm)polyacylamide (6%) gels in TAE or TBE
to separete DNA fragment of about 500bps (from some RT-PCRs);
I've see that there are in some catalogues precast horizontal PAGE, often I
nead more lanes or i nead more long lanes, I don't have the supports for
more big vertical PAGE, but i've the thank for big (20x20cm) agarose gels:
Is it possible to use the horizontal thank of agarose gels to assemle an
horizontal PAGE? Can it be used for an horizontal electrophoresis?
Must be submerged from the TAE or not?
I never read something about this tipe of horizontal PAGE probably becouse
this methods don't work, but in that case why don't work?

When i do vertical PAGE in TAE usualy I don't have the problem of a staking
gel before running gel, i do only the "running" gel in the simple TAE 1X. 
Lately i nead to separete with PAGE some genomic dsRNA of a virus (IPNV) and
for do that i read a paper of Ganga et all. (Polyacylamide gel
electophoresis of viral genomic RNA as a diagnostic method for infectious
pancreatic necrosis virus detection-  Journal of Virological Methods, 50
(1994) 227-236) that suggest to use the classical methos of Laemmli
(Nature, vol 227-1970) and Studier (J.Mol.Biol. 1973-79,237-248).
That classical methods, used to separete proteins, use a stacking and a
running gel with differnt  concentration of acryllamide and different
buffers; why use that methods for dsRNA and not the more simple TAE or TBE?
Mybe the big concentration of dsRNA make necessary a stacking gel like in
the proteins?

This dsRNA of about 2000-2500 bps i suppose that run in the 6% PAGE in the
native condition but i haven't idea of how quick it run: i suppose more
than DNA but how much? Are there some information of the run of dsRNA in
the PAGE somewhere?
Thanks !


-- 
Fabio
FabioDB#deleteme#@katamail.com




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