Dwindling Plasmid yield *HELP*

Martin Leach leach at mbcrr.harvard.edu
Mon Apr 25 22:56:09 EST 1994

In article <2pi0biINNbqs at medicine.wustl.edu>, HAVILAND at KIDS.WUSTL.EDU
(David L. Haviland, Ph.D.) wrote:

> Martin:
> This may not apply to your situation but we have had something like this 
> before in our lab.  The cause turned out to be some sort of phage 
> contamination of our bugs that contained our eukaryotic expression vector 
> (with cDNA) of choice.  Do you notice any "stringy-stuff" accumulating 
> inyour cultures with overnight incubation?  This would indicate 
> lysed bacteria.
> In our case, our yeild of plasmid kept dropping until we were only getting 
> a few hundred micrograms from 2 liters of TBroth, and with each successive 
> plasmid prep the mess of lysed bacteria seemed to increase.  It goes 
> without saying that we had to do successive plasmid preps as didn't get the 
> yield needed the last time we did it, ergo - we furthered the infection 
> until the 'sharp-eye'd' tech in the lab looked at the bugs and said "That 
> looks like a phage infection!"  I should note that she's our local expert 
> in phage preps as well... grin!

Very interesting. In fact i store the 500ml of LB-broth in a cold-room that
has some old and decaying library. (in the form of large plates with
plaques on). They are pretty cruddy - fungus etc... but as ever dont like
to throw anything away. Will start cleaning the cold room.

p.s. whats the best way to clean bottles which may have phage gunk stuck on
the outside.

p.p.s. havent noticed the stringy stuff, will look harder (only certain
constructs though)



.....          Martin Leach                Email:leach at mbcrr.harvard.edu 
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