Screening large ccc plasmids

GRGGTA at PATERSON.CHRISTIE-HOSPITAL.MANCHESTER.AC.UK GRGGTA at PATERSON.CHRISTIE-HOSPITAL.MANCHESTER.AC.UK
Mon Jul 26 04:16:00 EST 1993



Does anyone have a reliable method for the isolation and electrophoretic
separation of large (50-300 kb) ccc plasmids from bacteria?  We would like
to screen a culture collection consisting of environmental strains of
Vibrio and Pseudomonas spp. for large plasmids.

Russ Herwig
Department of Microbiology SC-42
University of Washington
Seattle, WA  98195

Several years ago I used a method to screen large (up to 200kb I think,
maybe more) ccc plasmids from a series of Pseudomonad plant pathogens
which was developed for the purpose of differentiating between strains
with some success. This involved scraping some bugs from a plate or using
a broth culture and lysing them in situ in the wells of a vertical agarose
gel using lysozyme and/SDS in very gentle fashion. With some practice
reliable plasmid patterns were produced reproducably. I cannot get the 
published references on this right now, but could get them if you are 
interested. A. Malik et al in the Journal of General Microbiology in the 
early 1980's would be informative.
Hope this is some use
Graham Atherton



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