Abstract: Using the screening medium containing X-gal, 10 strains with high producing lactase activity were isolated from 10 yak yogurt cheese samples purchased from Xizang. An efficient lactase producing strain RTM-111 rescreened out from the 10 strains was identified as Bacillus subtilis by the means of morphological feature, physiological and biochemical characteristics measurement and 16S rDNA sequence analysis. Lactase enzyme properties research showed that the optimal reaction temperature and pH were 50℃ and 7.5. The enzyme was stable at 40~50℃ and pH6.5~8.0. The lactase activity was promoted by Na⁺, Mg²⁺ and Mn²⁺, and strongly inhibited by Cu²⁺, Zn²⁺ and EDTA.The high producing activity of B. subtilis RTM-111and the stability of the lactase made it potentially useful.