Study On Diversity Of Slow-growing Rhizobiaisolated From Vigna Radiata

Studies on diversity and phylogeny of slow-growing rhizobia isolated from Vigna radiata in China was conducted by using phynotype,16S rDNA gene PCR-RFLP , 16S rDNA gene sequencing, 16S-23S rDNA IGS PCR-RFLP and RAPD assays.Results of phenotype test shown that all 44 rhizobia tested and 7 reference strains were clustered into three groups(I,II.III) which were furthered divided into subgroups at the similarity of 78%. Group I include subgroup 1 and 2 which contained 13 slow-growing rhizobia isolated from Lianyungang, Shijiazhuang and Handan. Group II consisted of Subgroup 3 and Subgroup 4.The former included strains isolated from Liangyungang, Handan and B. japonicumUSDA110 and the later included strais from Xinjiang and Sichuan and reference strain B. Haoningense2281.Group III was composed of strains isolated from Guangdong , Guangxi and type strains of B. elkanii USDA76, USDA46 and USDA86.Results of 16S rDNA gene PCR RFLP analysis revealed that all strains tested could be clustered into three groups at the similarity of 76%. Group I contained of 13 slow-growing rhizobia including LYG1.Group II consisted of 21 strains tested and type strains of B.japonicum and B.liaoningense.Ten strains isolated from Guangdong ,Guangxi and the type strain of B. elkanii.composed of Group III.Results of 16S-23S rDNA IGS PCR-RFLP shown that strains tested could be divided into A and B groups, which could be correspondently subdivided into AI, AII, AIII, BI and BII subgroups at the similarity of 85%. Compared with 16S rDNA PCR-RFLP, IGS RFLP assay shown higher resolution, All strains tested could be divided into 21 IGS RFLP patterns. A significantly geographical effect on biodiversity was demonstrated from strains isolated from Xinjiang, Guangdong and Guangxi regions.Results of RAPD analysis shown that all strains tested could be divided into a, b, c, d, f and g subgroups with higher resolution and similarity of 89%.Comparing with the results of phenotype, 16S rDNA gene PCR-RFLP , 16S rDNA gene sequencing, 16S-23S rDNA IGS PCR-RFLP, RAPD analysis revealed higher similarities and reflected the effect of geographical factors on genetic biodiversity of rhizobia.