16S rDNA Genotyping of Lactic Acid Bacteria Using PCR-RFLP Analysis

Abstract

The 16S rDNA genotypes among lactic acid bacteria were determined using PCR-RFLP analysis. Eight tetrameric restriction enzymes (AccII, AfaI, HaeIII, HhaI, MboI, MspI, TaqI, XspI) were used for RFLP analysis and adequate numbers of informative band were obtained from each enzyme. Eighty-two genotypes were obtained from 99 lactic acid bacteria. In the case of only two restriction enzymes, sixty-five genotypes, which are most in all combination, were obtained from the combination of XspI and MspI restriction patterns. Therein, fifty-two species could be assigned to specific genotype, supporting the application of this analysis for identification of bacterial species. The genotype assigned by PCR-RFLP using XspI and MspI showed 92.3% successful phylogenetic affiliations. PCR-RFLP analysis using 20 lactic acid bacteria was demonstrated, and the results corresponded approximately to the calculated fragment patterns. These results indicate that PCR-RFLP should be useful for genotyping, identification and approximately classification of lactic acid bacteria.