a couple things

Warren Gallin wgallin at gpu.srv.ualberta.ca
Thu Apr 21 10:21:53 EST 1994


In Article <sticknbd-210494102158 at 160.129.125.9>,
sticknbd at ctrvax.vanderbilt.edu (Henry) wrote:
>1  Does anybody know the molecular mechanism of DEPC
>inactivation of RNases?
>2  I had some trouble with RNA gels I was running
>recently.  It seemed there was a variable amount of
>precipitation in the wells, and very little 18 & 28S.
>Swapping out several solutions (formaldehyde with
>some precipitation, yellowed 10X mops) made no
>difference.  What solved it was changing my EtBr.
>The stuff I had been using was stored bench top at
>room T.  The stuff I just started using was from the
>same batch (I made it up in October 1993) but stored
>refrigerated.  The bench top stuff was somewhat
>more opaque and definitely darker than the refrigerated
>aliquots.  Anybody know what went on with the EtBr
>and why/how it ruined my gels?  I never heard of EtBr
>going bad.


1) DEPC covalentlyu reacts with active site residues in the RNase, this
irreversibly inactivating it.
2) If you have a solution of anything that is cloudy after having been left
at R.T., one good possibility is that something grew in it.  Mind you,
anything that grows in EtBr is one weird organism.  Anyway, that would
explain the precipitation and probably the problem with your RNA; the
creature produced RNase that chewed your RNA.  This is a speculation, and is
not based on experience or authority.


Warren Gallin,
Department of Zoology, University of Alberta
wgallin at gpu.srv.ualberta.ca



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