Differentiation of soybean-nodulatingBradyrhizobiumUSDA 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.