Engineering Deletions in the Mg1-Pk1 Gene Cluster of Streptomyces sp. Mg1 to Abolish Production of the Mg1-Pk1 Metabolite

Abstract

Many species of bacteria produce secondary metabolites in response to competitor species and changes to their local environment. Streptomyces sp. Mg1 (S. Mg1) produces the polyketide metabolite Mg1-Pk1, which participates in the defense of S. Mg1 when challenged with Bacillus subtilis. When the biosynthetic genes of the Mg1-Pk1 gene cluster are deleted, the resulting mutant strain, S. Mg1-Δ37, is hypersensitive to growth inhibition by B. subtilis. However, deletion of the Mg1-Pk1 biosynthetic genes required removal of ~80 kb from the gene cluster, a perturbation too large for genetic complementation. We aim to introduce a deletion in a single open reading frame that abolishes production of the Mg1-Pk1 metabolite and complement it for rigorous genetic analysis. In preparation, we have constructed and are currently screening a cosmid library in E. coli into which gene deletions will be engineered using the lambda red recombineering system and introduced into Streptomyces sp. Mg1 through homologous double recombination.