ethidium staining of RNA

mmccorm at macc.wisc.edu mmccorm at macc.wisc.edu
Sat Jun 24 18:35:48 EST 1995


In article <40.14305.3148 at channel1.com>, harold.drabkin at channel1.com
(Harold Drabkin) wrote:

> Subject: ethidium staining of RNA gels
> 
> K>interested in any information or recommendations and in what
> experiences K>other labs have had with the effect of ethidium staining on
> transfer of K>RNA from agarose gels. Is there another stain which would
> allow staining K>of the RNA with no effect on transfer.
> 
> My experience with electrobloting RNAs from polyacrylamide gels (10-15%,
> 7M urea (tRNA sized molecules)) is that I have no problems.  I do a
> quick stain in transfer buffer with EtBr, take my picture, then
> transfer (I'm using a TAE based transfer buffer).  Transfer time is
> usually two hours at 40 volts.
> I've also seen the recommendations against staining with EtBr; don't
> know if I get away with it due to the urea (which comes out during the
> staining process), acrylamide, or the small size of the RNA that I'm
> interested in.

I've had good luck with Sybr Green II from Molecular Probes. They claim
10-fold more
sensitivity than EtBr (without background fluorescence) and also that RNA
can subsequently blotted.
Takes about 30min to stain agarose gels with no destaining required. May
have to use a higher conc. of dye than recommended for formaldehyde gels,
but urea acrylamide gels are a piece of cake. I have no connection or
financial interest in Molecular Probes, I'm just happy with the product.

Good Luck.



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