[Histonet] what do people for ISH
mikael.niku <@t> helsinki.fi
Fri Jul 6 02:50:19 CDT 2007
Hello Jennifer, Emily, and others,
I think it's really best to be as careful as possible when one is
starting RNA work. But if the lab has been working succesfully for
years, I don't see why they should change the practices. Only if they
are now looking at very rare RNAs, requiring top quality materials for
maximal sensitivity, this might be relevant.
DEPC treatment of solutions is in my experience unnecessary, if one uses
high quality materials. So one can do without this highly toxic stuff.
MilliQ water is as such RNAse free (I tested ours by incubating my RNA
probes a few days at room temp... no detectable degradation). Using
RNAse free glassware & tools, RNAse free chemicals, and of course
gloves, the solutions will be RNAse free without DEPC.
The endogenous RNAses are probably really the most serious threat to
RNA. Tissues like pancreas are a nightmare, due to high enzyme content.
So after killing the animal, you need to work quickly and use effective
RNAse inhibitors / inactivators. We don't have separate processors,
microtomes, or cryotomes for RNA work, and I'm fairly satisfied with the
With best regards, Mikael
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