help needed with homogenizing

Eric R. Hugo ehugo at one.net
Thu Mar 20 22:38:07 EST 2003


Dear Navinisha,
    	Our lab processes a lot of adipose tissue which is about as sticky as 
you can get.  The best method in our hands has been grinding in LN2 
followed by exrtaction with TRI Reagent.  For the grinding step we find 
that it is essential to keep LN2 in the mortar at all times and to use LN2 
to actually rinse the powdered tissue out of the mortar and into a tube.  
The only trick with this procedure is keeping your own tissue from 
freezing.  Hope this helps.
Eric


Navinisha.Chetty at vcp.monash.edu.au (Navinisha N Chetty) wrote in 
news:14c42b014c2143.14c214314c42b0 at mail1.monash.edu.au:

> Hi there!
> I was wondering if anyone could help with a problem I am encountering 
> when trying to extract RNA from rat gut tissue.  At the moment I am 
> placing 30mg of gut tissue into lysis-binding solution (from the Ambion 
> RNAqueous-4PCR kit) and cutting it into tiny bits in an eppendorf tube, 
> with a pair of scissors, followed by homogenizing with a plastic pestle 
> provided in the kit.  I have found that I get very low yeilds of RNA 
> because the filter is always clogging during the isolation of RNA.  I 
> have also used the liquid nitrogen method with a mortar and pestle but 
> as I am using such small amounts of tissue, I loose much of the powder 
> when it melts on the mortar and pestle and thus again I get low yeilds 
> of RNA.  The gut tissue is very mucousy and stringy and I do not know 
> of any other methods to obtain a high yeild of RNA from a small amount 
> of tissue.  Can anybody please help me???       
> Thanks,
> Nav
> 
> ---
> 




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