[Q] How to purify small PCR frg. ?
popa0206 at PO-Box.McGill.CA
Wed Jul 19 17:58:20 EST 1995
> p_rudolph at MORPH.SPACENET.DE (Peter Rudolph) writes:
> Hi netters,
> two fragments: 29 and 67bp. She separated them by gel
> electrophoresis on a 2.5% AG.
: How to purify the bigger fragment from the
> gel slice with a really *high* yield?
> Any suggestions?
> Any help will be highly appreciated!
The only thing I can think of Electroelution or DEAE-paper.....
Another thing you could do is use a two vector cloning method...
You have the vector you want to clone in say with amp R and then
a second vector with TetR....
You clone the fragment (96 bp) into the tet R plasmid
cut with your restriction enzyme ligate to the othe amp R plasmid
then select for amp R/TetR
then cut with two enzymes one near your fragment and another neat where the
two vector sequences are ligated together such that your "DNA" is in only the
AmpR half of your plasmid then plate and select for ampR.
If this is hard to follow (as I know it must be) there is a reference In Biotechniques a
while back (Jan 1995 or Dec 1995)
Graham Dellaire Snail Mail:
Red Cross, Research
McGill University Montreal Blood Services
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