Mutation in thecobOgene generates auxotrophy for cobalamin and methionine and impairs the symbiotic properties ofSinorhizobium frediiHH103 with soybean and other legumes

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We report here the isolation of a methionine and cobalamin mutant strain (SVQ336) of Sinorhizobium fredii HH103 obtained by Tn 5-lacZ mutagenesis. Sequence analysis showed that the transposon was inserted into a gene homologous to cobO. This gene codes for a cobalamin adenosyltransferase which is involved in the biosynthesis of vitamin B12. Another HH103 cobO mutant (strain SVQ524), was constructed by the insertion of Ω interposon. Both cobO mutants required the addition of methionine because cobalamin acts as a cofactor of the enzyme MetH, which catalyses the last step of the methionine biosynthesis. Mutant SVQ524 failed to nodulate on Vigna radiate but was able to nodulate on Glycine max cvs. Williams and Peking and Cajanus cajan, although the total number of nodules formed was highly reduced in comparison with that of plants inoculated with the wild-type strain HH103. The roots of these plants did not seem to secrete enough cobalamin and/or methionine to support growth of cobalamin/methionine auxotrophs in the rhizosphere. In all cases, the phenotype of SVQ524 was nearly overcome by the addition of methionine or cobalamin to the plant growth media or by the presence of a copy of the cobO gene in cosmid pMUS756.

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