RNA Denaturation on PAGE

Warren watchcity at my-deja.com
Tue Feb 1 21:57:40 EST 2000


Hello, I presume when you say you are concerned about rna "denaturation", you
mean degradation, or are you concerned they are renaturing?  I have run a lot
of labelled transcript purification gels, and maintaining a gel temp of
almost painful to touch (using aluminum heat diffusion plates to alleviate
"smile" and thermal cracking) with the formamide denaturation conditions you
have described worked well.  I always preran the gels for ~ 30 minutes before
loading.  I think having "fresh" formamide soln. helps.  The conditions
recommended for dna sequencing work well.  Finally, I never saw any
appreciable amounts of the smaller bands that would indicate degradation.  I
was taught that freeze-thaw was the worst thing for rna.  good luck,  Warren

In article <877690$h88$5 at pegasus.csx.cam.ac.uk>,
  "Choona" <xuna68 at hotmail.com> wrote:
> Hi,
>
> Should polyacrylamide gels containing 7M urea be run at elevated temp to
> ensure denaturating conditions throughout? Should the gel be pre-heated? I
> heat my samples in formamide buffer to 70, but worry my RNA is denaturing
> whilst running.
> Please reply to cb252 at mole.bio.cam.ac.uk as I've borrowed a friend's
> account.
> Thanks so so much!!!!
> Claire Balding
>
>


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