The invention relates to a method for increasing content of docosahexaenoic acid (DHA) in schizochytrium limacinum grease. The method is characterized by comprising the following steps: lowering metabolic flux of a pentose phosphate pathway in thallus by selecting a fermentable carbon source, and lowering the metabolic flux of the pentose phosphate pathway in thallus by further adding an exogenous regulation factor; inhibiting the activity of malic enzyme in a tricarboxylic acid transfer system to greatly lower the content of nicotinamide adenine dinucleotide phosphate (NADPH) in the schizochytrium limacinum thaluus, promote the large-scale synthesis of DHA and enable the content of DHA in the grease of the obtained thallus to be as high as 55.3%, wherein the fermentation carbon source is 1,2-propylene glycol or any one or a mixture of 1,2-propylene glycol and acetate, the exogenous regulation factor is preferably sesamol which is added after filtering and degerming when a fermentation culture medium is sterilized, and the addition amount of the sesamol is 0.5-3.0mM/L. The method disclosed by the invention is simple and easy to perform, obvious in effect, capable of greatly increasing the content of DHA in the thallus grease and easy for industrial application.