On 6 Dec 1995, Shei-Pon Tzung wrote:
> Hi all:
>> What is the best commercially available poly(A) RNA isolation kit?
> I have tried Promega and Qiagen, which gave me a yield about 0.2 to 0.6
> %? I have been struggling with this for months, Please help.
>> Shie-Pon Tzung
> Fred Hutchinson Cancer Center
> E-mail address: stzung at fred.fhcrc.org>>
Moving out of the 'kit-based' mindset would be a first step, I believe.
The good old oligo-dT cellulose (or sepharose, although I prefer the
former), in a 1-2 mL column made out of sterile 3 mL syringe and glass-wool
has always worked with excellent recovery. For a complete step-by-step
protocol, consult any molecular biology manual - the basic principle and
the steps are same in all the cases, following a clean total RNA prep.
One may have to standardize the conditions, like temp., buffer conc.,
whether to include SDS etc., depending on the source organism and the
abundance of the message of interest etc., but that should not take more
than a a couple of days.
Hiranya S. Roychowdhury
Plant Genetic Engineering Lab.
Box 3GL, NM State Univ.
Las Cruces, NM 88003
Phone: (505) 646-5785
hroychow at nmsu.edu