Inserting a large DNA fragment into a large plasmid

Paul Jasper pjasper at orion.luc.edu
Thu Sep 16 11:56:41 EST 1999


Don't worry too much about getting large DNA fragments into E. coli.
We transform 25kb plasmids into E. coli (by electroporation).... and it
works just fine.  The transformation efficiency will be lower than
normal but you should have no problem getting colonies.

We've tried up to 50kb plasmids with success.  The only other thing I
would suggest is to be very careful about growing conditions... Do not
let the bugs overgrow!  (I usually go 12 hours) ... When the bugs
overgrow, weird things happen, such as deletions and aggregation.


paul

-- 
___   ____   ____   ____   ____________________   ____   ____   ____   _
\  \ / \  \ / \  \ / \  \ /    Paul Jasper     \ / \  \ / \  \ / \  \ /
/\  \  /\  \  /\  \  /\   Micro/Immuno - Loyola U. /\  \  /\  \  /\  \
  \__\/  \__\/  \__\/  \__pjasper at orion.luc.edu__\/  \__\/  \__\/  \__\/



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