Genetic analysis of a novel plasmid pZMX101 from Halorubrum saccharovorum: determination of the minimal replicon and comparison with the related haloarchaeal plasmid pSCM201

The DNA sequence of a novel haloarchaeal plasmid pZMX101 (3918 bp) from Halorubrum saccharovorum was determined and six ORFs were predicted. The largest ORF encodes a putative replication initiation protein RepA, which shares 40% sequence similarity with the Rep201 of a theta-replication plasmid pSCM201 recently isolated from Haloarcula, suggesting that pZMX101 might replicate via a theta-type mechanism. Using pZMX101 as the only haloarchaeal replicon, a shuttle vector pZMX108 was constructed and successfully transformed into Haloferax volcanii DS70. Based on this in vivo system, the minimal replicon (1978 bp) of pZMX101 was determined. It is composed of the repA gene plus c. 400-bp upstream and 300-bp downstream sequences. Significantly, the putative replication origin of pZMX101 and that of pSCM201 contain different types of sequence motifs, and these two plasmids exhibit distinct host preference for Haloferax and Haloarcula, respectively.