In order to significantly increase the yield of mupirocin to meet production demand,the cultivation conditions of the fluorescent Pseudomonas aeruginosa strain MF21-25,w hich produces mupirocin,including colony characteristics,fermentation process,and fermentation raw materials,were optimized .Through the optimization of culture conditions to obtain the best culture conditions of the high-yield strain and the key influencing factors of shake-flask fermentation .Mupirocin productivity under optimum fermentation conditions was 6693μg/mL in shake flask culture,which was 52 .2% higher than that under the original condition (4398 μg/mL) . The research results can provide basic theoretical and technical support for pilot-scale amplification and industrial production .