Transformation of Streptomyces erythraeus

Streptomyces erythraeus strains were transformed efficiently with six different plasmid DNA vectors by a protocol that uses 0.2 mM Ca2+, 5 mM Mg2+ and 10% DMSO to enhance the stability of protoplasts to storage at -80 degrees C and their transformability at 30 degrees C. The primary thiostrepton-resistant (Thior) transformants for most vectors were unstable even when grown selectively. This instability did not appear to be due to incompatibility with indigenous S. erythraeus plasmids. Conversely, stable transformation was not the result of plasmid or host mutations. Transformation instability or plasmid copy number thus prevented successful shotgun-cloning of DNA that complemented the eryD mutation, which blocks the biosynthesis of erythromycin A, because only plasmid DNA without an insert could be isolated from a Thior Ery+ clone.