Intergeneric conjugal transfer of plasmid DNA from Escherichia coli to Kitasatospora setae, a bafilomycin B1 producer

Abstract

An effective transformation procedure for Kitasatospora setae was established based on transconjugation from Escherichia coli ET12567 (pUZ8002) using a phi C31-derived integration vector, pSET152, containing oriT and attP fragments. While no transconjugation was observed under the standard transconjugation conditions for Streptomyces species, sufficient transconjugation (>1 x 10(-6)) was achieved on ISP4 medium containing 30 mM MgCl(2) using a 25- to 125-fold excess of E. coli donor cells. In addition, the sequence and location of the chromosomal integration site attB of K. setae was identified for the first time in genera of non- Streptomyces actinomycetes. K. setae contains a single phi C31 attB site. Similar to the case of Streptomyces species, the attB site of K. setae is present within an ORF encoding a pirin-homolog, but the K. setae-attB sequence deviates slightly from the consensus sequence of Streptomyces attB sequences.