Most Consistent Total RNA Extraction Method/Kit?

Eric Anderson e-anderson at ski.mskcc.org
Wed Nov 27 16:09:51 EST 1996


In article <mnoda-2711961014160001 at 207.126.101.77>, mnoda at lipovx.lbl.gov
(Michael Oda) wrote:

> Hi,
>      We are trying to assess the relative concentration of total RNA in
> liver tissue as one of the measures of the metabolic state. Currently
> we have tried a number of methods including the Guanidinium Isothiocynate-
> "Chomczynski method" and a RNA extraction kit from Qiagen. But due
> to the variability of the phenol extraction in the Chomczynski
> method and an unidentified variability with the Qiagen columns we never 
> got reliable results. So I was wondering if anyone out there has
> come across a reproducible method of extracting RNA. Any pointers
> would be welcomed.

michael,

we've gotten great results with both the CsCl protocol from Current
Protocols and by using Tri Reagent from Sigma.  the differences seem to be
that the CsCl protocol isolates more HMW RNA and is a little light on the
smaller stuff where as the Tri Reagent gives a more even distribution of
transcripts.

both methods give excellent quality, intact RNA.  the main differences are
in cost:  almost nothing for the CsCl if you already have the CsCl, the
tubes (and of course an ultracentrifuge and a rotor) compared to about
$165/200ml of the Tri Reagent (1ml will treat 50-100mg of tissue or
.5-1*10^7 cells); and of course speed.  i can do 4 samples in about 1.5-2
hours with the Tri Reagent (about 45 min. hands-on) vs. about 6-8 hours
for the same number of samples with the CsCl (about 2 hours hands-on
time).

either way, it's your choice.  both work fine in a number of hands in our
lab but all but the most retro member of the lab have switched to the Tri
Reagent for speed and ease of use.

good luck,

eric



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