zimprich at nefo.med.uni-muenchen.de
Sun Nov 28 12:24:44 EST 1999
In the last few months, i have a very big cloning problem,.
I try to get a AOC1 (5´overhang CC!TGAGG), ECO R1 (also 5´overhang
G!AATTC) fragment into a vector also cut with AOC1, ECO R1. It sounds
quite easy, but it is a nightmare.
I tried it over a dozen times, but i only get very few colonies after
transformation, which were all wrong. I did all kinds of control
experiments; so let us assume that Ligation, Transformation etc. do work
in my hands.
Is it possible the some restriction enzyme overhangs get lost, or need a
special treatment for ligation?
I purify my DNAs over agarose gels and clean them up using Quiagen
Thanks a lot
-------------- next part --------------
An HTML attachment was scrubbed...
More information about the Methods