In this study, four significant factors of seven were checked by one-way test and Plackett-Burman test using different carbon and nitrogen sources, pH, temperature, inoculum, rotational speed, and time as single variables. And the steepest-climbing test was carried out on the screened factors. The Box-Behnken test was used to investigate the optimal conditions for intracellular polysaccharide production by liquid fermentation of Poria cocos. The results showed that inoculum, pH, glucose mass fraction and yeast paste mass fraction were the key factors for the production of intracellular polysaccharides by liquid fermentation of Poria cocos bacteria. The content of intracellular polysaccharides in Poria cocos under the conditions of inoculum of 9%, pH 5.7, glucose mass fraction of 4%, yeast paste mass fraction of 1%, time of 7 d, temperature of 28 ℃ and rotational speed of 170 r/min. Intracellular polysaccharide was (5.82±0.18) g/L and biomass was (14.63±0.21) g/L, which were 1.6 and 2.0 times higher than those before optimization, respectively. Compared with conventional cultivation, the time for intracellular polysaccharide production by liquid fermentation of Poria cocos was greatly shortened, the consumption of pinewood was reduced, and the cost of Poria cocos liquid fermentation medium was inexpensive.