Medium compositions and fermentation conditions for bio-oil production by fermentation of corn straw hydrolysate with IFFI01367 strain were explored, and the lipid composition was identified. The optimal conditions were determined as follows: 5% sugar, 1% yeast extract as nitrogen source, initial pH 5.8, 30°C, 150 r/min, 50 mL corn straw hydrolysate in 300 mL erlenmeyer flask, fermentation time 4 d. Under these conditions, the biomass reached 23.23g /L, oil content was 42.32% and the oil production capacity was 9.83g /L. The main fatty acids of the oil induced palmitic acid, oleic acid and stearic acid; linoleic acid was more rich, which is convenient to post-processing.