Bonehead ligation driving me crazy

John Ladasky ladasky at
Fri Mar 15 08:16:09 EST 2002

ladasky at (John Ladasky) wrote in message news:<c09b237b.0203022248.1ab5db5f at>...

> > What about trying to gel purify your BamHI/AflII insert?  It might
> > perhaps take 40 minutes longer, but your assured of a clean and
> > correclty sized insert.
> I've just tried this, with the BamHI/BsrGI combination.  I got eight
> colonies this time, which I'm screening.  The yield is still extremely
> poor.  If I get my construct this time, great.  But I would sure like
> to understand the source of this problem before I have to do any more
> molecular biology.

Well, to follow up, all eight of my colonies were exactly what I
wanted.  I still need to know why my ligation and/or transformation
reactions have gone sour, but the immediate problem of obtaining my
molecule has been solved.

Thanks to every one who offered advice!

John J. Ladasky Jr., Ph.D.
Department of Biology
Johns Hopkins University
Baltimore, MD 21218

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