Differentiation of soybean-nodulating Bradyrhizobium USDA strains using restriction fragment length polymorphism analysis of 23S-5S rRNA genes

Cluster analysis of the electrophoresis patterns from polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) of approximately 3 kbp of the 23S-5S rRNA genes (rDNA) of 21 Bradyrhizobium USDA strains and a Sinorhizobium fredii type strain was conducted to determine the usefulness of these genes for differentiating soybean-nodulating bacteria. The PCR-RFLP analysis of 23S-5S rDNAs separately digested by each of the four restriction enzymes HhaI, HinfI, MboI, and XspI detected 20-26 restriction fragments longer than 100 bp and a total of 13 RFLP patterns in the Bradyrhizobium USDA strains tested, which were distinguishable from the S. fredii type strain. Thus, PCR-RFLP analysis of 23S-5S rDNAs is a simple and useful approach to the grouping of soybean-nodulating isolates, although there were topological differences between the results of PCR-RFLP analysis of these genes and of the 16S-23S rDNA intergenic transcribed spacer region. Depending on the design of the analysis, it is necessary to choose the appropriate target region (i.e. the 16S-23S rDNA ITS region or the 23S-5S rDNA) because of differences in the number of operational taxonomic units in the resolutions of each target beween Bradyrhizobium japonicum and Bradyrhizobium elkanii.