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Formaldehyde, hybridization, and Northern blots

SLF forsburg at sc2.salk.edu
Sun Jul 17 14:40:43 EST 1994


In article <Ct37I6.G4y at acsu.buffalo.edu>, camhuber at ubvms.cc.buffalo.edu
(Joel Huberman) wrote:

> It seems to me, however,
> that the formaldehyde-modified RNA should not be able to hybridize with
a probe
> after it has been Northern-blotted to a membrane.  Obviously, it does hybri-
> dize;  otherwise formaldehyde gels would not be used for Northern blottings.
> Thus, the formaldehyde reaction must be reversible.  I would like to know whe-
> ther the formaldehyde reaction is fully reversed under standard hybridization
> conditions, or is there a possibility that use of a formaldehyde gel leads to
> less efficient hybridization than would use of a standard gel or a different
> type of denaturing gel, such as a formamide gel?

Reading the GeneScreen manual and others, I have always been led to
believe that baking the filters after transfer is imperative with
formaldehyde blots to "reverse the formaldehyde reaction."  I have never
empirically determined whether this makes a difference.  I do use a lower
formaldehyde  concentration than usually called for although I believe
these days so does everyone else.

susan

-- 
forsburg at sc2.salk.edu
    formerly forsburg at molbiol.ox.ac.uk



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