Transposition and transduction of plasmid DNA in Streptomyces spp

Abstract

To expand the application of molecular genetics to many different streptomycete species, we have been developing two potentially widely applicable methodologies: transposon mutagenesis and plasmid transduction. We constructed three transposons from the Streptomyces lividans insertion sequence IS493. Tn5096 and Tn5097 contain an apramycin resistance gene inserted in different orientations between the two open reading frames of IS493. These transposons transpose from different plasmids into many different sites in the Streptomyces griseofuscus chromosome and into its resident linear plasmids. Tn5099 contains a promoterless xylE gene and a hygromycin-resistance gene inserted in IS493 close to one end. Tn5099 transposes in S. griseofuscus giving operon fusions in some cases that drive expression of the xylE gene product, catechol deoxygenase, giving yellow colonies in the presence of catechol. We have also developed plasmid vectors that can be transduced into many streptomycete species by bacteriophage FP43. We describe the characterization of FP43 and mapping of several bacteriophage functions. The region of cloned FP43 DNA essential for plasmid transduction includes the origin for headful packaging.