Stability of plasmid pA387 derivatives in Amycolatopsis mediterranei producing rifamycin.

Abstract

Genetic studies on the biosynthesis of rifamycins in producer strains such as Amylcolaptopsis mediterranei U-32 are severely hampered by the availability of efficient transformation procedures and stable plasmid vectors. Using an efficient electroporation procedure we have studied the replication and stability of a pA387 derivative, pDXM32. This plasmid confers enhanced plasmid stability and copy number compared to pA387 derivatives commonly used as cloning vectors in A. mediterranei. Deletion derivatives in the region previously identified as being a minimal replication origin were also examined with respect to their ability to transform A. mediterranei and at least one locus was essential for replication. A 5.4 kbp DNA fragment was sequenced and annotated encoding the replication and plasmid stability functions. A parA homologue was identified which is likely to confer plasmid stability.