RNase-inhibiting agents

Dr. Peter Gegenheimer PGegen at UKans.edu
Fri Nov 14 18:50:02 EST 1997

On Thu, 6 Nov 1997 11:07:10, Ralf Oltmanns <Biologist at SpyRing.com> 

> I am trying to extract mRNA from soil bacteria via direct lysis.
> Therefor I need a very potent RNase-inhibiting agent.
> I used GITC (guanidin-isothiocyanate) and DEPC (diethyl-pyrocarbonate)
> in different protocols with not really satisfactory results, whereas
> they really work fine with pure cultures.

A very convenient inhibitor is aurin tricarboxylic acid (ATA). It is
freely soluble at high concentrations (must be pH'd to neutrality; 
buy the ammonium salt). As I recall, one normally uses 5 to 10 mM, 
but even 20 mM should be OK. Like heparin, mentioned by another 
poster, ATA is difficult to remove from the RNA if you want to treat
the RNA enzymatically. (ATA can be separated, at least partially, on
a gel-filtration column, either Sephadex G10 to G25, or Biogel P 
series). The original reference is Chelm
& Hallick, about 1980-82, prob. NAR or Anal. Biochem. 
| Dr. Peter Gegenheimer       |  Vox: 785-864-3939  FAX: 
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