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blotting of large RNAs

Nick Theodorakis nicholas_theodorakis at urmc.rochester.edu
Thu Jan 20 11:02:21 EST 2000


In article <38872220.ABBB60DE at gmx.net>, Frederik
=?iso-8859-1?Q?B=F6rnke?= <ricky_boernke at gmx.net> wrote:
> Hi all,
> I'd would like to improve transfer of large RNAs during the
> blotting
> procedure. AFAIK incubation of the gel with NaOH or NH4AC prior to
> blotting should help. Has anybody a detailed protocol?


Soak in 50 mM NaOH for 30 min. (In real life, I only go 20 min., 'cause
the NaOH step makes me nervous)

Soak in 0.1M Tris pH 7.4 for 30 min.

Blot in 10X SSC
(I've been using charge-modified Nylon e.g., GeneScreen Plus or Biodyne
B, etc.)

Nick



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