Validaton of RNA integraty

Dr. David J. Meyer meyerdj at phibred.com
Thu Nov 19 09:49:28 EST 1998


mlush at hgmp.mrc.ac.uk (Mr. M.J. Lush) wrote:

>	Were producing total RNA to generate a cDNA library
>(we have too little tissue to extract mRNA:-(.  
>	Do we need to do a Northern blot to demonstrate that the
>RNA is intact or is visualizing the 18S and 28S rRNA bands on an 
>EtBr/agarose gel good enough evidence that that batch of RNA OK?

>TIA

>-- 

>Michael
>~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~
>NPC rights activist           |      Nameless Abominations are people too.

A useful method which consumes little RNA is to use your preparation
as template in an in vitro translation and examine the (radiolabeled)
products by SDS-PAGE. If you see translation products larger than,
say, 70 kDa you know that you have some translatable and fair-sized
mRNAs present. This takes less time than doing a northern blot. One
potential problem is that your RNA may contain contaminants which
interfere with translation (e.g. polysaccharides, vanadyl
ribonucleosides if you have used them) or could be modified by DEPC if
you have not taken care to remove it.

Good luck!
Dr. David J. Meyer
Quality Traits
Pioneer Hi-Bred International, Inc.

meyerdj at phibred.com
7300 NW 62nd Ave.
Johnston, IA   50131-1004

Ph. 515/254-2639
FAX 515/254-2619
Email: meyerdj at phibred.com




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