Screening And Identification Of Highly Efficient Soybean Rhizobia

Twenty Sinorhizobium fredii strains designated as from ZX01 to ZX30 and from QJ1 to QJ30 were isolated from soil samples from Chongqing by using pot plant tests. One hundred strains of S.fredii stored were revived. Their nitrogen fixation efficiency of total 160 S. fredii strains was compared by using pot plant test with soybean cultivars Ningzhenland Yudou 1. The mean dry weight per plant inoculated by strains SMH12, ZX20, HH29 and HW1 was 3.80, 3.64, 3.38 and 3.12g respectively, which were 179%, 167%, 148% and 129% higher than reference strain HN01, and higher than nitrogen fertilizer by 42%, 36%, 27% and 17%. SMH12 and ZX20 ranked the first and second. Then, taking NH4NO3, TA11 and HN01 as control, the secondary screening was done with the above four strains. ZX20 and SMH12 took the first and second places. The dry weights per plant inoculated with these two strains were higher than TA11 by 16.09% and 10.76% respectively. Statistical analysis results showed that the SMH12 and ZX20 maintained significantly higher nitrogen fixation ability than strain HN01 and TA11.The nitrogenase activities of strain HW1, HN01, SMH12, HH29, TA11 and ZX20 were 11.84, 10.56, 10.14, 6.58, 4.66, 3.81 umol C2H4 /g h respectively on Ningzhen 1 cultivar and 9.56, 2.29, 5.28, 1.84, 5.86 and 3.92 umol C2H4 /g h on Yudou 1 cultivar. No significant relationship was observed between nitrogen fixation ability and nitrogenase activity. The nitrogen content of the plants inoculated with SMH12, ZX20 and HN01, as well as the nitrogen content of blank control plant were 2.26%, 2.44%, 2.10%, 1.70% respectively. The nitrogen content of the plants inoculated with SMH12 and ZX20 were significantly higher than blank control and plants inoculated HN01.Antibiotics resistance, salt tolerance, huge plasmid profile, acid and alkaline resistance were determined too. It was found that both SMH12 and ZX20 were resistance to Neo and Amp among the 9 test antibiotics. SMH12 could grow at salt concentration 2.5% while ZX20 only at 1.5%. SMH12 could grow at pH value from 4.0 to 12.0 while ZX20 only from 4.0 to 10.0. Three and two huge plasmid were detected respectively from ZX20 and SMH12 by in-sity rapid plamid-detecting method.In order to determine the competitive nodulation abilities of SMH12 and ZX20, the two strains were marked with luxAB by introducing recombinant plamid pHN102 into the rhizobial cells. Assay of stability of the plamid showed that the plamid was inherited stabley in the cells. The results of plant sand pot experiment showed that both SMH12 and ZX20 presented lower competition abilities than HN01. The percentage of nodulation occupancy of SMH12 and ZX20 were 40% and 45% respectively. When SMH12 and ZX20 competed with indigenous rhizobia of experiment field in this school, the nodulation occupancy of these two strains was as low as 8% and 4% respectively. However, when competed with indigenous soybean rhizobia at Qijiang, SMH12 and ZX20 had a high nodulation occupancy percentafe of 60% and 80%Field test in Zhongxian County and Qijiang County showed that inoculating SMH12 and ZX20 significantly increased the yield of soybean. In Zhongxian County, the meanyield of soybean inoculating SMH12 was 4% higher than control and the yield inoculating ZX20 was 12% higher than control. In Qijiang county, inoculating SMH12 and ZX20 increased greatly the nodule numbers and nodule weights.