Discrimination of Rhizobium tropici and R. leguminosarum strains by PCR-specific amplification of 16S-23S rDNA spacer region fragments and denaturing gradient gel electrophoresis (DGGE)

Abstract

With the aim of detecting Rhizobium species directly in the environment, specific PCR primers for Rh. tropici and Rh. leguminosarum were designed on the basis of sequence analysis of 16S-23S rDNA spacer regions of several Rh. tropici, Rh. leguminosarum and Agrobacterium rhizogenes strains. Primer specificity was checked by comparison with available rDNA spacer sequences in databases, and by PCR using DNA from target and reference strains. Sequence polymorphisms of rDNA spacer fragments among strains of the same species were detected by denaturing gradient gel electrophoresis (DGGE). The specific PCR primers designed in this study could be applied to evaluate the diversity of Rh. tropici and Rh. leguminosarum by analysing the polymorphisms of 16S-23S spacer rDNA amplified from either whole-cell or soil-extracted DNA.