Abstract: The production of feruloyl esterase activity by Lactobacillus could be screened by assaying transparent zone on the selective agar plate and high efficiency liquid chromatography. The results of experiment showed that only Lactobacillus fermentation ATCC 9338 had feruloyl esterase activity, and the enzyme activity was 260mU/L. Shotgun cloning method was used to clone and sequence feruloyl esterase gene from this strain. This study will provide theoretical basis for feruloyl esterase-producing probiotics.