Characterization of a plasmid in isolates of Corynebacterium sepedonicum

Lysis of mid-log phase cells of the Gram-positive bacterium, Corynebacterium sepedonicum, by a combination of lysozyme treatment and incubation with alkaline sodium dodecyl sulfate at 56 degrees C led to the recovery of a single plasmid. The plasmid was purified in CsCl density gradients, and its molecular weight estimated to be 31 megadaltons (46 kilobases), as determined from its relative mobility in agarose gels, from its contour dimensions in electron micrographs, and from the size of the fragments generated when it was cleaved with various restriction endonucleases. Thirteen widely divergent isolates of C. sepedonicum were screened for the presence of plasmid, and of these, 11 were shown to harbour a single plasmid at a level of about 30 copies per cell. Cleavage of the plasmid with PstI gave an identical banding pattern in agarose gels for the fragments from all of the plasmid-carrying isolates. The relevance of plasmid incidence and distribution in C. sepedonicum is discussed in relation to the possibility of developing a test for the detection of bacterial ring rot by using plasmid DNA as a hybridization probe.