Abstract

Many of the bacterial genes involved in nodulation (nod) and nitrogen fixation (nif) are dispersed over the 500-kilobase plasmid pNGR234a of the broad host-range Rhizobium species NGR234. As a first step toward generating a complete physical and genetic map of the plasmid, a full overlapping collection of cosmids was derived from a total genomic library. Clones were aligned by combining fingerprinting, hybridization, and pulsed-field gel electrophoresis data. Symbiotic loci were localized by probing a representative set of cosmids with both homologous and heterologous genes. nodABC, nodD1, nodD2, nodSU, nolB, and region II are widely dispersed over pNGR234a, while the two functional copies of nifKDH are separated by only 28 kilobases. Interestingly, sequences homologous to nodE, nodG, nodP, and nodQ have been assigned to another autonomously replicating element in Rhizobium species NGR234. Similarly one copy of the structural dctA gene is located on the symbiotic plasmid (dctA1) while the other is on what we assume to be the chromosome.