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Method for selection of transposable DNA and characterization of a new insertion sequence, IS493, from Streptomyces lividans.

Department of Molecular Genetics, Lilly Research Laboratories, Eli Lilly and Company, Indianapolis, Indiana 46285.

ABSTRACT

A method to select for transposable elements from Streptomyces spp. by using insertional inactivation of a repressor gene that functions in Escherichia coli was developed. Plasmid pCZA126, which can replicate in Streptomyces spp. or E. coli, contains a gene coding for the lambda cI857 repressor and a gene, under repressor control, coding for apramycin resistance. E. coli cells containing the plasmid are apramycin sensitive but become apramycin resistant if the cI857 repressor gene is disrupted. Plasmids propagated in Streptomyces spp. can be screened for transposable elements that have disrupted the cI857 gene by transforming E. coli cells to apramycin resistance. This method was used to isolate a new 1.6-kilobase insertion sequence, IS493, from Streptomyces lividans CT2. IS493 duplicated host DNA at the target site, had inverted repeats at its ends, and contained two tandem open reading frames on each strand. IS493 was present in three copies in the same genomic locations in several S. lividans strains. Two of the copies appeared to be present in regions of similar DNA context that extended at least 11.5 kilobases. Several other Streptomyces spp. did not appear to contain copies of IS493.

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