RNA preps - looking for suggestions

theodorn at medlib.georgetown.edu theodorn at medlib.georgetown.edu
Tue Jun 9 09:12:28 EST 1998


In article <bpmurray*STUFFER*-0806982145540001 at macmac-2.ucsf.edu>,
  bpmurray*STUFFER*@socrates.ucsf.edu (Bernard Murray) wrote:
>
> In article <357C8CAB.292C at science.uottawa.ca>, mbeyers at science.uottawa.ca wrote:
>
> > Theo Thijs wrote:
> > >
> > > If you don't care about the RNA yield why don't you try TriZol buffer?
> > >
> >
> > Well, actually, I do care about the yield and I've tried Trizol and such
> > in the past.  Recalling that a perfect RNA sample should have an
> > A260/A280 ratio of about 2.0, I've done no better than achieve 1.4 with
> > Trizol.  However, the prep I'm using ALWAYS gives me 2.0 +/- 0.02.
> >
> > So my conclusion is that, to save two extra pipetting steps, people are
> > paying huge amounts of money for the convenience of Trizol.
>
> I don't think home-made Trizol is that expensive (you're just paying
> for the guanidinium and phenol mainly).  I agree that the ratios can
> be suspiciously low at times but the RNA looks good on a gel and
> probed bands are generally nice and sharp and this is what counts.
>      I also agree that one method does not necessarily fit all (although
> Trizol comes close) so by all means stick to what works.
>      For high speed spins (eg. up to 20 k rpm) in Sorvall or Beckman rotors
> I use plastic (polythene?) tubes from Sarstedt and these are nuclease
> free and phenol resistant.
>      Bernard
> --
> Bernard Murray, PhD
> Dept. Cell. Mol. Pharmacol., UCSF, San Francisco, USA
>

Okay, I lost track of the original article, but I thought the original post
was regarding the GITC/acid phenol method, which is basically Trizol, without
whatever modification is put in to recover DNA.  In any case, "switching" to
Trizol will not solve the problem.  The trick is to change volumes or spin
speed.	If I am purifying RNA from small > <input type=

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