Bacitracin high-producing strains were bred by low energy plasma implantation mutagenesis and the fermentation condition was optimized with bacillus licheniformis as original strain. When the mutagenesis time was 15 s, the lethal rate of the strain was 80%, and the positive mutagenesis rate was 38%. After several rounds of breeding, a high-yield mutant strain C15 was obtained, and the yield reached 638.47 U/mL, which was 38.89% higher than the original strain. The optimum conditions of fermentation were determined via single factor and orthogonal experiments: the carbon source was 40.0 g/L corn starch, nitrogen source was 80.0 g/L soybean cake powder, and the temperature, initial pH and rotating speed was 37 ℃, 7.0 and 280 r/min, respectively. Under the optimum conditions, the yield reached 880.87 U/mL. The verification test was carried out in a 50 L fermenter, and the final cumulative bacitracin yield of the mutant strain reached 927.36 U/mL. Low-energy plasma implantation mutagenesis, as a new and efficient microbial breeding method, has a wide prospect in the field of strain improvement.