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fermentation of coli and plasmid stability

S.Ballal ballal_sv at hotmail.com
Tue May 13 01:08:52 EST 2003

Dear Ede,

 I may sound stupid but are you really losing plasmids (I mean
experimentally proven or your are just worried about the possibility.)

With my(though limited) experience with E.coli with a set up similar
to yours(industry with expression of protein), we actually have
observed very little or no plasmid loss after fermentation in absence
of selection pressure. Though I know it differs from one expression
system to other. I am not sure it is a very common phenomenon with
high copy no. plasmids.  EK or DUNCAN  to comment please.

Some people feel that high growth rate causes loss of plasmids as they
are not able to partition properly in very fast growing cells. Though
I am not sure about it's implication in high copy no. plasmids. 
Reducing the growth rate in such a case should obviously be tried.

Anyway discussion continues

Indus Biotherapeutics Ltd.
Ahmedabad, India

Ede <bggfg at gmx.de> wrote in message news:<vjqvbv89ai20ch4h2hv718jts9duopstbl at 4ax.com>...
> On Mon, 12 May 2003 10:10:08 +0100, Duncan Clark <junk@[]>
> wrote:
> >Historians believe that in newspost 
> ><3EBB385C.6080700 at biotech.NOSPAM.ntnu.no> on Fri, 9 May 2003, Trond Erik 
> >Vee Aune <trondaun at biotech.NOSPAM.ntnu.no> penned the following literary 
> >masterpiece:
> >>You could try using some kind of partition (par) or killer system (i.e. 
> >>hok/sok)
> >
> >I agree fully. Look for a paper in Biotechnology Vol 6 December 1988 p 
> >1402-1405 by Kenn Gerdes with hok/sok on a small 580bp cassette. 
> >Stabilisation factor of 1000 for pBR322 and 10000 for pACYC, p15 ori, 
> >derivatives
> >
> >Duncan
> I`m working in a biotech company and when i use this hok/sok system it
> have to be patent free. somebody have informations 
> More about my plasmid: Its a high copy plasmid with an kanamycin
> cassette, but in an industrial process you cannot use antibiotics, and
> this plasmid is to express proteins (whatelse) but the thing is that
> plasmdi free cells overgrow cells with plasmids. So the process is
> shit, but i have to show at the end of the fermentation that most of
> the cells still have the plasmid!!! I hope somebody have an idea
> Ed

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