Ligations

[JOHN NASH]

Hi there,

MN>I have been suddenly having the utmost frustrating experiences with ligations


May I ask a few questions??  I need more information.

1.  What is the replicon of the plasmid?  Will it replicate in the host you are
transforming?  What is the host in question?  Which method of transformation
are you attempting?

2.  Can you transform uncut plasmid?  What is your selection for transformants?
Does the selection require "expression" before plating?

3.  What amounts of DNA are you trying to ligate and transform?  What
restriction enzyme are you using?  Can you detect ligation on a mini-gel?

These questions may seem a bit trivial, but to a reader at the other end of
the message (like me), it makes a difference between speculation and a more
educated guess (is there a difference ;-) ?? )


     cheers,
     John,

--------------------------------------------------------------
     John H.E. Nash <Bitnet: NUM208JN at NRCCAD.NRC.CA >
     Institute for Biological Sciences,
     National Research Council of Canada,
     Ottawa, Canada  K1A 0R6.

     Phone:  (613) 990-0990     Fax:    (613) 952-9092.
==============================================================