The nir, nor, and nos denitrification genes are dispersed over the Bradyrhizobium japonicum chromosome

Cleavage of genomic DNA from Bradyrhizobium japonicum strain 3I1b110 by the restriction enzymes PmeI, PacI, and SwaI has been used together with pulse d-field gel electrophoresis and Southern hybridization to locate the nirK, norCBQD, and nosRZDFYLX denitrification genes on the chromosomal map of B. japonicum strain 110spc4. Mutant strains GRK13, GRC131, and GRZ25 were obta ined by insertion of plasmid pUC4-KlXX-aphII-PSP, which carries recognition sites for the enzymes PacI, PmeI and SwaI, into the B. japonicum 3I1b110 n irK, norC and nosZ genes, respectively. Restriction of strain 3I1b110 genom ic DNA with PacI, PmeI and SwaI yielded three, five and nine fragments, res pectively. Pulsed-field gel electrophoresis of restricted mutant DNAs resul ted in an altered fragment pattern that allowed determination of the positi on of the selected genes. Complementary mapping data were obtained by hybri dization using digoxigenin-labeled B. japonicum 3I1b110 nirK, norBQD and no sZD as gene probes. The nirK, norCBQD and nosRZDFYLX genes were located clo se to the groEL(2), cycH and cycVWX genes, respectively, on the strain 110s pc4 genetic map. In contrast to other denitrifiers, B. japonicum 3I1b110 de nitrification genes were dispersed over the entire chromosome.