Transformation of plasmid DNA into Streptomyces at high frequency

Abstract

OVER 60% of known antibiotics¹ are produced by Streptomyces species, including many substances with valuable clinical and other applications; they therefore have considerable medical, biological and commercial importance. Although the process of gene exchange mediated by conjugation within these actinomycetes is apparently widespread², the transfer of genetic material between individuals by such sexual means is, by definition, predominantly restricted to members of the same species. Transfer of genes as free DNA (transformation) or by viral mediation (transduction) has been well documented in some microorganisms^3–5, but the efficiency of these processes also falls dramatically with increasing divergence of the species involved, as the homology of their DNA sequences decreases. In any case, the occurrence of these processes in streptomycetes remains to be established. This situation has hitherto precluded the generation of new combinations of DNA sequences from diverse actinomycete species which could have profoundly beneficial effects on both the range of chemical structure and the quantity of antibiotics produced⁶. We report here the development of a plasmid transformation system for Streptomyces which should allow the cloning of any DNA sequence into these organisms and which, potentially, provides a means of directly manipulating the pathways of antibiotic production. The system involves the uptake of covalently closed circular (ccc) DNA by protoplasts in the presence of polyethylene glycol (PEG) and the visual detection of transformants, at high resolution, after regeneration of the protoplasts.