ligation problems and electroporation

theveno at cc.umanitoba.ca theveno at cc.umanitoba.ca
Fri Oct 6 12:05:04 EST 1995


In article <450p4g$nnp at edisto.awod.com>, bkodrzyc at awod.com (Bob Kodrzycki)
wrote:

> We are experiencing ligation problems with "routine" ligations.  I
following this
> newsgroup I see that we are not the only ones.
etc.

Bob;  are you sure the HMW stuff you see is what you want? It could be
concatamers not supercoiled or circular DNA hence a low transformation
eficiency.  Concatamers could be favoured if your DNA conc is to high in
your ligation rx. You may have to fool around with your DNA conc and
ratios in your ligations.

Ultracompetent cells are just as good or at least close to
electrocompetent cells nowadays so I wouldn't worry to much about the
cells being the problem. I admit I almost exclusively use electroporation
as we were lucky enough to score a machine from a lab that hardly ever
uses it. Once you have a stock of cells made and stored at -70°C it is a
little easier and less time consuming than the chemical method. The price
of cuvettes is obscene (what isn't from these companies) so we reuse them
like most other people.

Dave Boyd
University of Manitoba

-- 
Hey, hey, hey, hey! It was the DNA.
Hey, hey, hey, hey! That made me this way.



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