Abstract: Using paddy soil as test material, the author screened and separated herbicidal fungi with the method of plate dilution combined with bioassay of target plants. The results showed that 11 out of 32 strains had higher inhibitory rates (>50%) on the root length and plant height of lettuce. 5 out of the 11 strains had higher inhibitory rates (>40%) on the plant height of barnyard grass, however, 3 strains had inhibition and 2 strains had no inhibition on rice growth by bioassay. The strain No.8, which had higher inhibition on growth of barnyard grass but was harmless on rice, was selected for study of the growth curve. The strain No.8 was identified as fungus Aspergillus by form and molecular biological identification. The optimization of cultivation conditions for the strain No.8 were using sucrose as carbon source, yeast extract or peptone as protein source and at 30℃, pH 6. The ferments of strain No.8 contained 5.14 mg/L proteins, 5.59 mg/L phenolic compounds, 448.12 mg/L soluble sugar and 2.39 mg/L flavonoids. The results demonstrated that the strain No.8 could be used in paddy field weeding.