Two self-transmissible Sym(biosis) plasrhids, one encoding pea-specific nodulation and nitrogen-fixation functions (plasmid pJB5JI) and the other encoding clover-specific nodulation and nitrogen-fixation functions (plasmid pBRlAN) were used to determine whether the symbiotic genes encoded on these plasmids are expressed in various members of the Rhizobiaceae. The host specificity of Rhizobium trifolii and R. leguminosarum Sym plasmid-cured strains could be directly determined by the transfer to these strains of the appropriate Sym plasmid. The nodulation of white clovers was restored by either plasmid pJB5JI or pBRlAN when these plasmids were transferred to two transposon TnS-induced hair-curoing (Hac-) R. trifolii mutants. In addition, lucerne nodulation was testored to a Hac-R. meliloti mutant when either plasmid pBRlAN or pJB5JI was transferred to this strain. The phenotype of nonmucoid (Muc-) Rhizobium mutants, which had altered cell surfaces, was not influenced by the transfer to these strains of plasmid pBRlAN or plasmid pJBSJI.

Non-nodulating mutants of Rhizobium trifolii were found in cultures incubated at 35 °C for 7 days. From among ten strains incubated at the elevated temperature six became non-nodulating. The frequency of Nod~ mutants in R. trifolii cultures varied from 1 to 75 % depending on the strain tested. It is suggested that gene(s) controlling nodulation are located on a large plasmid.

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A conjugal plasmid which encodes both pea nodulation genes and nitrogenase genes, and which is labeled with the transposon TnS, was transferred to a wildtype Rhizobium trifolii strain to examine the stability and expression of the host range and fixation (Fix') phenotypes. Transconjugants were isolated which were shown to initially form nitrogen-fixing nodules (Nod' Fix') on both clovers and peas. These hybrid strains were then repeatedly passaged through either pea or clover nodules or onto a solid agar medium to determine whether these broadened-host-range characteristics were stably maintained. An instability was noted in the capacity of some of these hybrids to form nitrogen-fixing nodules on all of the host plants used. The broadened nodulation ability was, however, more on August 4, 2020 by guest

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