RNA electrophoresis!!

Regina Shaw regina at biotech.ufl.edu
Mon Nov 3 09:22:43 EST 1997


In article <345A61CC.5290 at hotmail.com>, Han <huangz at hotmail.com> wrote:

> Dear netters,
> 
> I encountered a problem in RNA formaldehyde electrophoresis: I added EB 
> in my formaldehyde agarose gel(final concentration 50ug/100ml) and run 
> gel for 2 hours. Amazingly, when I visualize gel under UV lamp I found 
> very high staining background which evenly distributed and composed 
> almost 2/3 area of gel starting from the margin near the wells. This high 
> flurescent staining is so strong that overhided my RNA bands. I would be 
> very appreciate any suggestion about the possible reasons. My RNA 
> formaldehyde electrophoresis buffers are as following: (please save me 
> out of the nightmare)
> 
Hi -
We recently followed New England Biolab's suggestion to do RNA
electrophoresis in native TBE/agarose gels, with good results. We added
ethidium bromide both to the gel and to the TBE electrophoresis buffer, to
a final concentration of 0.5 microgram per ml each. RNA samples were
heated for 3 min at 65 degrees C in denaturing sample buffer, loaded
immediately and run for about 1.5 hours. We got good separation of RNA
bands and very little background. See BioTechniques vol. 9, no. 5 (1990),
p. 558-560.
(no affiliations with NEB except for being a satisfied customer)
Hope this helps,
Cheers,
Regina
Recombinant Protein Expression Lab/Molecular Services Core/ICBR
Gainesville, FL, USA



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