Abstract: The present invention discloses a method of improving the fermentation yield of polyunsaturated fatty acids, in which Schizochytrium limacinum is used as production strain to produce polyunsaturated fatty acids (PUFA), and glycine betaine or trehalose is added to the fermentation medium. In the present invention, after Schizochytrium limacinum fermentation system is treated with exogenous glycine betaine, the yield of PUFA produced by fermentation with Schizochytrium limacinum can be greatly improved. The present invention significantly improves the yield of PUFA produced by microorganism, reduces the cost without harming the environment, and saves manpower and material resources by simple and effective regulation of fermentation, and thus the method is simple, convenient and cost-effective.