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Novel densitometric method for endonuclease analysis of Escherichia coli DNA samples containing multiple plasmids.

Medical Microbiology Laboratory, Funabashi Medical Center, Chiba, Japan.

ABSTRACT

We developed a novel method whereby the digestion pattern of each plasmid was distinguished in a sample of endonuclease-digested plasmid DNAs which contained multiple plasmids extracted from a bacterial strain. This method consisted of two procedures. (i) The concentration ratio of each undigested plasmid DNA and of each digested DNA fragment was calculated on the basis of densitometric scanning of an electrophoretogram, and the concentration ratio was then compared with the theoretical concentration ratio to determine from which plasmid each fragment was generated. (ii) The second procedure involved rapid visual identification with a scanning graph. We thus analyzed five strains of Escherichia coli that harbored several cryptic plasmids. The two procedures made it possible to analyze the restriction digestion pattern of each plasmid without the need to isolate the individual plasmids, except in the case of certain fragments. Even when the digested patterns of these exceptional fragments could not be distinguished completely, however, our method had the advantage that peak patterns of plasmids could be compared visually among different bacterial strains.