BETTER METHOD THAN: Freeze & Squeeze: DNA from Low-melt

David Friedman dfried at mendel.genetics.washington.edu
Tue Oct 29 13:28:20 EST 1991


In article <1991Oct23.233111.18225 at agate.berkeley.edu> nigel at codon1.berkeley.edu (Nigel Walker) writes:
>He has described to me a technique which he calls freeze & squeeze, for 
>isolating DNA from low-melt agarose.  The excized band is diluted in buffer 1
>[this is where the cookbook is important!], frozen with liq N2, and centrifuged
>through a tiny hole in the bottom of a 0.5ml eppendorf tube (into a 1.5ml tube)

Many of the responses to this post have included protocols using extractions
and glass wool.  We have had much success recently with a much simpler
protocol adapted from a blurb in the April 1991 TIG (p. 119?).  Instead of 
glass wool, you use a piece of Whatman filter paper.  If done correctly, 
the paper prevents the agarose from entering the 'eluate'.  There is no 
need for freezing or extraction.  We use a very high grade agarose to 
minimize impurities, and get excellent yields from TBE gels.  The resulting 
eluate can be directly added to a ligation mixture.  This proceedure has 
saved me gobs of time and is practically effortless.  I highly recommend 
giving it a try.  
--
_______________________________________________________________________________
David Friedman                          
Dept. of Genetics, SK-50             "I am not a number,
U of Washington, Seattle WA 98195          I am a Friedman!"



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