nothern problems

kim at m44.unm.edu kim at m44.unm.edu
Mon Mar 18 13:42:58 EST 1991


In article <9103181117.AA01788 at genbank.bio.net>, bourson at jouy.inra.fr (Claude Bourson) writes:
>Northern blot on nylon membrane or nitrocellulose membrane.
>
>I have problems to transfert RNA on membrane after using Glyoxal/DMSO
>denaturing agarose electrophoresis.system.
>RNA seems to have good electrophoresis behaviour, but I am not able to
>hybridize it after transfert on Nylon membrane (Amersham Hybond N+).
>If you are familiar with northern after Glyoxal/DMSO electrophoresis,I would gre
>   atly appreciate to have more explanation on the techniques you use, and more
>details on the type of membrane (nitrocellulose, nylon ...) which works well.

As a guess, I am wondering if the glyoxal adducts on the RNA are interfering
with hybridization.  These adducts have to be removed from the RNA after
electrophoresis by alkali treatment before blotting (I think pH 8 is
necessary), otherwise, they continue to perform their function of preventing
base pairing.
Daniel Kim



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