Stabilization of Streptomyces lividans by Homologous Recombinational Insertion

Abstract

We have developed a system for the introduction and maintenance of novel tandem repeats in the chromosome of Streptomyces lividans 66. This was achieved by introducing, via transformation, Escherichia coli “suicide” vectors carrying manipulated S. lividans DNA fragments. Selection for antibiotic resistance markers carried on such plasmids permitted the isolation and maintenance of mutant strains containing novel tandem repeats formed by the integration into the chromosome of the plasmids, via homologous recombination between plasmid–borne chromosomal sequences and identical sequences on the chromosome. When novel repeats were introduced, and maintained, in regions of the chromosome which become deleted in unstable strains of S. lividans, those deletion events were blocked. Surprisingly, such strains were also 10 to 20–fold more stable than the parent even in the absence of selection. In stable regions of the chromosome, the maintenance of novel repeats had no obvious effect on the deletion events. This strategy could be generally applicable to industrial strains of Streptomyces, where instability is a common problem.