Characterization of soybean rhizobia at different levels using PCR based techniques
采用PCR-RFLP技术在不同水平上鉴定大豆根瘤菌

Nineteen standard USDA strains representing three recognized soybean rhizobia,Sinorhizobium fredii ,Bradyrhizobium japonicum and B. elkanii, were examined by restriction fragment length poymorphism (RFLP)analysis of 16S rRNA genes amplified by polymerase chain reaction (PCR). Four composite genotypes were obtainedfrom the combined data of the RFLP analysis with three endonucleases. S. fredii, B. japonicum, B. elkanii II andIIa reference strains fell into thase four genotypes respectively. No genotype was shared by two species. Therefore,16SrDNA PCR-RFLP is a rapid tool for the idenification of soybean rhizobia.According to 16S rDNA PCRRFLP, 22 fast-growing and 19 slow-growing soybean rhizobia strains from China were identified to S. fredii andB. japonicum, respectively.The reference strains and wild type isolates were the examined by 16S~ 23S intergenic spacer PCR-RFLPanalysis. PCR results indicated that the length of the intergenic region between S. fredii and Bradyrhizobium sp.is different. All fast-growing S. fredii strains produced one larger band (2. 1 kb), and all slow-growing Bradyrhizobium strains produced one smaller band (2.0 kb). Based on 16S~23S RFLP analysis , 22 S. fredii isolates canbe further separted into two "genotypes" that originated from two different regions in China; 19 B. japonicumstrains from Heilongjiang Province (northeast of China) can be separated into 11 genotypes. There wild type isolateswere also characterized at strain level by REP (repetitive extragenic palindromic) and ERIC (enterobacterial repetitive intergenic consensus) PCR fingerprinting.