DNA extraction of 140 kb piece from agarose

Alan Lyons lyons at cellt.demon.co.uk
Fri Nov 10 06:58:19 EST 1995


In article <msz-1011951240160001 at image.bmc.uu.se>, msz at bio.embnet.se
(Michael Szardenings) wrote:

> In article <DHJDrs.4n9 at freenet.carleton.ca>, bm335 at FreeNet.Carleton.CA
> (Christina Chan) wrote:
> 
> > I have been having difficulty in isolating a 140 kb piece of DNA
> > from a TBE agarose (1% SeaKem GTG) plug.  The plug contains
> > ~500 ng of DNA.  I have tried the Gene Clean II kit but was un-
> > successful.  
> > 
> > Any suggestions?
> If it is 140 bp, then you will really have problems with most silica based
> kits. Use a kit designed for PCR fragment purification. I used 'Jetpure'
> from Genomed and they guarantee, that you loose 100% of all primers etc
> below 50 bp. Thus the yield in your case might not even be the best with
> such kits, but it worked well for me in most cases.
> Besides, you might be more successful with electroelution, its the ideal size.
> Good luck
>       Michael

Another thing you might like to try is to use your Gene Clean II kit (with
a TAE gel), and do your incubation with glass milk at 55 degrees (instead
of 4). This has worked for me for 200bp fragments. You also might like to
try Bio 101 mermaid kit, but don't use TBE, use TAE. 
Good luck, 
        Susan.

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