Localization of symbiotic mutations in Rhizobium meliloti

A total of 5 Nod- and 57 Fix- symbiotic mutants of Rhizobium meliloti strain 41 have been isolated after either nitrosoguanidine or Tn5 transposition mutagenesis. Chromosomal locations of mutations in 1 Nod- and 11 Fix- derivatives were ascertained by transferring the chromosome (mobilized by plasmid R68.45), in eight fragments, into symbiotically effective recipients and testing the recombinants for symbiotic phenotype. Alternatively, the kanamycin resistance marker of Tn5 was mapped. In five mutants the fix alleles were localized on different chromosomal regions, but six other fix mutations and one nod mutation tested did not map onto the chromosome. It was shown that the chromosome-mobilizing ability (Cma+) of R68.45 was not involved in the mobilization of genes located extrachromosomally. Moreover, Cma- derivatives of R68.45 could mobilize regions of the indigenous plasmid pRme41b but not chromosomal genes. Thus, mobilization of a marker by Cma- R68.45 indicates its extrachromosomal location. With a 32P-labeled DNA fragment carrying Tn5 as a hybridization probe, it was shown that in five extrachromosomally located Tn5-induced fix mutants and one nod mutant Tn5 was localized on plasmid pRme41b. This is in agreement with the genetic mapping data.