Efficient gene transfer of high copy number mibilizable plasmids containing a marineRhodobacter origin of replication

The replication region of pRD31, a 3.1-kb endogenous plasmid from a marineRhodobacter species, has recently been localized and sequenced. We report here incorporation of this replicon into a narrow-host-range mobilizable pBR325 derivative. This has allowed us to establish an efficient conjugative gene transfer system for a hydrogen-producing marine species ofRhodobacter that is unable to grow aerobically. Efficient transfer was obtained (1.2×10^–3 transconjugants per recipient cell), and hybrid plasmids replicated with a high copy number (>10) and good stability. Southern hybridization analysis indicated that the new vectors, pRDP203 and pRDP203s, were maintained in marineRhodobacter sp. NKPB0021 as autonomous replicons without detrimental structural rearrangements, confirming their suitability for use as shuttle vectors.