RNA resuspension problem

NJ njuni at poppy.ocn.ne.jp
Thu Jun 21 06:39:23 EST 2001


The gel-like substance may be polysaccharides.
The same thing sometimes happens to me when the amount of starting material
is too much as against the volume of RNA extraction system.

2 possible solutions to remove polysaccharides from the pellet:
1) Second round of RNA purification from the pellet: For example, add TRIZOL
to the pellet (or suspension), and proceed to homogenization and extraction.

2) LiCl (w/o alcohol) precipitation: Resuspend the pellet in 2M ("Molecular
Cloning" says 0.8M) LiCl with pestle or pipetting, and centrifuge at 4C.
Large RNAs (mRNAs) precipitate, while polysaccharides and small RNAs (tRNAs)
remain soluble. 
--
NJ

in article 20010619150239.15314.qmail at ww02.jatek.com, Melanie at
mmhierl at yahoo.com wrote on 6/19/01 3:02 PM:

> I have in the past had no problems precipitationg and resuspending my RNA in
> DEPC H20.  However, the last time I tried to do this using 1/10 volume NaAc
> ph 5.5 and 2 volumes EtOH and freezing at -20 overnight, I come up with a
> clear gel like blob of a pellet that I can't get to redissolve.  I have
> heated at 65 for 10 min several times, vortexed, flicked the tubes
> repeatedly, etc.  Should I heat the tubes longer?  Will that harm the RNA?
> Any other suggestions?
> thanks
> 
> 
> <http://www.biowww.net/forum/read.php?f=1&i=2567&t=2567>
> 




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