The endophytic fungus BS002 with good effect of disease-resistant has been isolated from the medicinal Sophora flavescens. The conditions of the fermentation medium were explored in this paper. On the basis of the single factor experiments, CH3COONa, potato and glucose were defined to be the main factors by Plackett-Burman Design; Response surface methodology (RSM) with Box-Behnke Design was used to optimize fermentation conditions of endophytic fungi BS002. The optimal conditions are defined as follows: 100 mL in the container 250mL, potato 246.47 g/L, glucose 27.81 g/L, CH3COONa 1.95 g/L, concentration of fungus 107 cfu/ml, temperature 25 oC, yeast extract 1.0 g/L, speed 150 rpm, fermentation for 4 d, dry weight of mycelium can reach 10.398 g/L.