Molecular characterization of symbiotic root nodulating rhizobia isolated from lentil (Lens culinaris medik.)

Rhizobia strains were characterized using molecular techniques along with traditional techniques such as physiological, biochemical and intrinsic antibiotic resistance. Two molecular techniques based on PCR amplification such as repetitive extra genomic palindromic (REP-PCR) sequences and restriction fragment length polymorphism (RFLP) analyses were used. Groupings generated by PCR DNA finger printing with either extragenomic palindromic repetitive primers or two different primers were correlated with similar levels of resolution. Although less discriminating, PCR-RFLP analysis of intergenic genes coding for 16S r-RNA yielded intra-specific monomorphisms indicating that six strains were closely related. The classification of strains was independent in agreement with those obtained by REP-PCR DNA fingerprinting. On the basis of Box profiles, five genotypes were identified among six strains. Nitrogen fixation nature of all strains was confirmed by amplification of nifH gene and no variations were observed among the strains. Variations in 16S r-RNA between Rhizobium leguminosarum and Bangladeshi strains observed by RFLP which revealed that Bangladeshi lentil symbiont might be different from Rhizobium leguminosarum. Moreover, physiological and biochemical test showed that these strains had more survivable capacity under stress conditions and in-vitro experiment indicated that they were highly effective in nitrogen fixation than the control as well as TAL-638 strain.