Question on Plasmid Cloning

A.F. Simpson AFS7 at le.ac.uk
Tue Aug 17 18:38:47 EST 1999


no one wrote:
> 
> These are called concatamers and they are a problem.  They will ligate into
> the vector just fine and the colonies will grow just fine.  The only way you
> can spot them is to do a plasmid prep on each colony and run that on a gel.
> If concatamers are there, they will run slower on the gel (i.e. the insert
> will have a higher molecular weight).

And to be even more sure, do a restriction digest with two different
enzymes which lie on either side of your insert (and don't cut the
insert).  Don't just stick to sites in the MCS either.  For example, the
PvuII sites in Bluescript are very handy for that, becuse they give a
600pb fragment in empty vector, so you can be sure the digest worked.

> Gary Krause
> Wayne State University

love
Anna



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