slow growing cultures

MUNGO at UVVM.UVIC.CA MUNGO at UVVM.UVIC.CA
Thu Aug 25 11:26:42 EST 1994


Perhaps someone could give me an explanation, and a solution for a problem
I have with a particular plasmid.  This plasmid is bluescript witha cDNA
insert aprox. 2.5 Kb long.  It was retrieved from a lambda ZAP II library
using the exassist phage.  As phage this clone grew well, however once it
was isolated as a plasmid it grows very slowly.  My impression is that every
time I subculture it the growth is even slower (this may be paranoia...).  I
can accept the theory that the insert encodes something toxic to the bacteria
and that there is leaky expression from the plasmid.  If this was true why any
growth at all ?  If it was toxic wouldn't the bacteria either die or kick it
out ?  Is it possible to have a sort of fading culture like I describe where
it gets weaker and weaker all the time ?  Would changing the bacterial host
strain help ?  The plasmid is presently in Stratagenes SOLR strain as that
is what I get back from the excision procedure (i know nothing of this strain).
I have clones very similar to this one (same family of genes) that grow fine
and I cannot see this clone being toxic.  Anyone have any insights or
suggestions ?
Thanks
Mungo



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