TROPLIST: FW: RNA purification for microarray analysis

[Dawid, Igor (NIH/NICHD) [E]]

Jose,

 

Below is a comment from a colleague in our lab. I hope it is useful.

 

Igor

Igor B. Dawid 
Laboratory of Molecular Genetics 
NICHD, NIH 
Bethesda MD 20892 
301-496-4448 or -7940 

________________________________

From: Tanegashima, Kosuke (NIH/NICHD) [F] 
Sent: Tuesday, November 27, 2007 2:01 PM
To: Dawid, Igor (NIH/NICHD) [E]
Subject: Re: TROPLIST: RNA purification for microarray analysis

 

Hi Igor

I purified total RNA with Trizol followed by Rnasey colomn purification.
It seems to me that the combination is critical for removing yolk. Also
I did not treat with Dnase: the combination of two methods is enough for
removing most of DNA from sample, and DNAse treatment sometimes ruins
the sample.

Kosuke

Kosuke Tanegashima
LMG, NICHD, NIH
Bldg 6B, Rm 420
Bethesda MD 20892
301-496-9689



________________________________

From: Jose Luis Mullor [mailto:jmullor@cipf.es] 
Sent: Tuesday, November 27, 2007 1:12 PM
To: troplist@list.mail.virginia.edu
Subject: TROPLIST: RNA purification for microarray analysis



Dear Troppers,
we'd like to do some microarrays on trops embryos, st. 12.5, but we had
a hard time passing the quality control from the Affymetrix team.  In
short, our RNA samples degrade and are not of sufficient quality to give
consistent results. The RNA Integrity Number (RIN) for our samples goes
from 7 to 8.5, but never higher. We tried several purification methods
and kits and collection protocols and still can't break through the 8.5.
Here is a sample of how they look on a gel:


I would appreciate it if anybody could send us some clues, tips,
protocols, etc. 
Best,
Jose L. Mullor

 

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