eluation of small DNA fragments from gel

Johns053 at mc.duke.edu Johns053 at mc.duke.edu
Wed Feb 23 12:26:20 EST 2000


I do this fairly routinely using the EluTrap from S&S.  If you want a low tech
version of this, do the old-fashioned electroelution into a dialysis bag using a
low molecular weight cut-off bag.




Matthias Wahl <matthias.wahl at gsf.de> on 02/23/2000 12:12:37 PM

To:   methods at hgmp.mrc.ac.uk
cc:
Subject:  eluation of small DNA fragments from gel


Could anyone tell me a good method for eluting small DNA fragments
(20-30bp) from Polyacrylamide? It could also be a method for eluting
from Agarose (Metaphor etc). My problem is, that most methods use
temperatures, at which such small fragments will melt.

I currently use 12% PAGE, and elute just by incubating together with
buffer for >2h at 37
-------------- next part --------------

?C and afterwards just filter away the
polyacrylamide. My problem is that I only have a very low efficiency in
a subsequent ligation step. Propably I carry over some contamination
from the polyacrylamide. Is this likely that remaining polyacrylamide is
inhibiting ligation reactions, or do I just have some impure reagents
for making the gel.

I also looked for some commertial kits for purifying the eluted DNA but
they are not suitable fur fragments of this size.

Thanks in advance

Matthias Wahl





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