Abstract: Abstract: The biogas fermentation is easy to fail resulting from high sensibility of methanogens to environmental change. So how to ensure the activity of methanogens is the key for anaerobic fermentation stability. A methanogenic microbial inoculant was constructed based on the physiological, biochemical characteristics and antagonisms of strain RY3, SH4, G1, G2 and G3. The performances of the inoculant under different pH values, temperatures and application rates of the methanogenic microbial inoculant were evaluated. It showed that the 5 strains had different physiological and biochemical characteristics as well as complementary roles. There were no antagonisms among 5 strains. The methanogenic microbial inoculant grew at pH value 5.5-10.5. Methane productions were 1 706.7-2 026.7 ?mols at pH value 5.5-9.5 after 3 days' culture, that of different pH values showed no significant difference respectively. The results indicated that the methanogenic microbial inoculant is resistant to acid and alkali changes. The inoculant grew at 15-70℃ and methane productions were 1906.9- 2028 ?mols at 30-60℃ after 3 days' culture, that of different temperatures treatment showed no significant difference respectively. The results indicated that the methanogenic microbial inoculant is adapted to a wide temperature range. At 20℃, total biogas yield of treatments 2%, 5%, 10% before 14 day were 234, 422 and 950 mL, and the methane concentration of treatments 2%, 5%, 10% on the 14th day were 46.9%, 51.2% and 58.9% respectively corresponding the treatments with 2%, 5% and 10% inoculation dosages of the methanogenic microbial inoculant. At 50℃, total biogas yield before 14 d were 2728, 3291 and 3 832 mL and the methane concentration on 14th day were 62.7%, 63.1% and 63.8% respectively corresponding the treatments with 2%, 5% and 10% inoculation dosages of the methanogenic microbial inoculant. The results indicated that the inoculation could shorten the starting time for methane production compared to the controls without inoculant at 20 and 50℃. Pilot tests by kinetic analysis indicated that inoculant could still shorten the starting time for methane production compared to the control using anaerobic active sludge as inoculant. At 20℃, total biogas and methane yields of 21 day of the treatment with 10% inoculation dosages of the methanogenic microbial inoculant were both 1.6 times of the control group with 10% inoculation dosages of anaerobic active sludge. At 50℃, total biogas and methane yields of 21 d of the treatment with 10% inoculation dosages of the methanogenic microbial inoculant were 2.7 and 2.8 times of control group with 10% inoculation dosages of anaerobic active sludge, respectively. Overall, this study showed that the methanogenic microbial inoculant could significantly improve the efficiency of biogas production at low temperature and high temperature, and it will provide a new technical way for optimal control on biogas fermentation.