Abstract: In order to obtain oil degrading mixed agents and optimized degradation conditions, sixty-eight bacteria, fifteen fungus and six actinomycetes were isolated from oily soil, oily wastewater, crude oil. Y-4 & Y-12 combination and Y-4 & Y-37 combination were selected through screening, rescreening and two-two mixed antagoinstic experiment, the oil degradation rates of the two mixed microbial agents were quite high by more than 60%. At the same time, the influence of different conditions on oil degradation of the mixed bacteria agents was studied. The results showed that, the degradation rates of the two mixed microbial agents, Y-4 & Y-12 combination in the initial pH 8.0, speed 160 r/min, the seed solution 0.5 mL with the ratio of 1:1, 30~ 35℃for 7 days, and Y-4 & Y-37 combination in the initial pH 7.5, speed 130-160 r/min, the seed solution 0.5 mL with the ratio of 1:1, 35℃for 7 days, were more higher with adding less initial crude oil volume and abundant matrix nutrition. The article experiment showed that, the oil degradation rates of the two mixed microbial agents were quite high under the appropriate conditions, and the degradation rate of the mixed microbial agents was higher than the single agent.