A functional map of the replicator region of the octopine Ti plasmid

Abstract

Using in vitro-generated deletion mutants of the octopine plasmid pTiB6, we localized a region specifying copy control and plasmid stability ( stb) on the right side of the replication origin ( ori). A region that prevents the establishment of nopaline Ti plasmids in strains carrying an octopine Ti plasmid was found to be situated to the left of the replication origin. This function appeared to be unrelated to plasmid incompatibility and has been called ein (for establishment inhibition). Strains harboring an octopine Ti plasmid from which the ein region had been deleted were good recipients for nopaline Ti plasmids. The ein − octopine Ti plasmids were themselves susceptible to establishment inhibition not only by nopaline Ti plasmids, but also by octopine Ti plasmids. Thus, octopine Ti plasmid derivatives that are ein − failed to establish—after introduction by transformation—in recipients carrying an octopine Ti plasmid. Rare transformants that did arise contained cointegrates between the residing octopine plasmid and the incoming ein − plasmid. Integration had taken place at, or close to the replication origin and did not affect virulence of the residing plasmid. After cointegration of an octopine Ti plasmid with an incoming ein − stb − plasmid, the resultant cointegrate showed the selective “amplification” of the internal ein − stb − plasmid during growth of the host in the presence of carbenicillin.