Bonehead ligation driving me crazy

[John Ladasky]

ladasky at my-deja.com (John Ladasky) wrote in message news:<c09b237b.0203022248.1ab5db5f at posting.google.com>...

> > 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
USA
Earth