RNA gel electrophoresis problems

cyc at glink.net.hk cyc at glink.net.hk
Mon Jun 9 12:45:02 EST 1997

On Mon, 9 Jun 1997, Lori Bentsen wrote:

> I have recently been having problems with my RNA gel electrophoresis. 
> In our lab we run radioactive RNA samples  ( labelled with P-32) on
> denaturing gels with the folloing ingredients:  31.5g urea, 7.5 mL TBE
> (tris, boric acid, EDTA) and 9.9 mL 38:2 % Acrylimide:Bis in the final
> volume of 75mL water.  To polymerize we add 375 uL Ammoinium persulfate
> and 37.5uL TEMED.  This gel has not given me any problems for almost 3
> years until now.  After the gel is finished running we transfer the gel
> to nitrocellulose and the gel has radioactive spots all over it.  This
> is making it very difficult to see the bands.  Please help!!!!!
> Lori Bentsen

Firstly,  as you are obviously running a PAGE then I don't see why you
don't just dry the gel on to Whatman paper and then do an autoradiogram as
this will save you the transfer time as well as all the mess one has to do
when doing nuclei acid transfers.  This also has the added advantage that
you will be looking at the actual gel rather than a tranferred version.

As for the high background radioactive spots,  I have never encounter this
sort of problems in my PAGE.  Have you checked your X-ray cassettes for
radioactive contamination? 

I presume you have re-run the PAGE to rule out a freak result.


C.Y. Cheung
University of Hong Kong.

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