Sequences essential for replication of plasmid pIJ101 in Streptomyces lividans

A 2.1-kb SacII DNA fragment of the high-copy-number Streptomyces lividans plasmid pIJ101 previously has been shown to include the functions required for maintenance of pIJ101 as an extrachromosomal replicon. This fragment contains the open translational reading frames rep and orf 56 plus an intervening segment believed to be noncoding. Using deletion mutations, we show that the pIJ101 replication origin and other cis-acting sites necessary and sufficient for replication map to a DNA segment that extends 515 bp 5' to the translational start of Rep and lacks orf56. Plasmids that include this segment are maintained in S. lividans as extrachromosomal replicons when complemented in trans by the rep gene product.