Screening large ccc plasmids

Paul N Hengen pnh at
Tue Jul 27 10:59:58 EST 1993

In article <1993Jul26.081619.19352 at>
>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.

>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 

Some early references are:

Eckhardt, T. 1978. A rapid method for the identification of plasmid
deoxyribonucleic acid. Plasmid 1:584-588.

Adachi, T. and V. N. Iyer. 1980. A procedure for the isolation and
purification of plasmid DNA from Rhizobium meliloti. Analytical Biochem.

Kado, C. I. and S. T. Liu. 1981. Rapid procedure for detection and
isolation of large and small plasmids. J. Bacteriol. 145:1365-1373.

Paul N. Hengen
National Cancer Institute
Frederick Cancer Research and Development Center
Frederick, Maryland 21702-1201 USA

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