A green process and environmentally benign process is highly desirable in the development of enzymatic catalysis. In this work, the shuttle plasmid pHA01 was constructed and the sucrose isomerase (SIase) was expressed in Bacillus subtilis WB800. The optimal nitrogen and carbon sources for SIase expression were yeast extract (15g/L) and un-pretreated cane molasses (UCM, 20g/L), respectively. After the UCM fed, the whole cell activity reached 5.2U/mL in a 7.5L fermentor. Optimum catalytic temperature and pH of whole cell were 35°C and 5.5, respectively. Although the biologic membrane reactor (BMR) system consecutively worked for 12 batches, the sucrose conversion remained higher than 90%, indicating the BMR system had a greater operational stability. Furthermore, isomaltulose production using the BMR system with low-cost cane molasses as its substrate not only reduces the production cost and mediates environmental pollution, but also solves the genetic background problem of the non-food-grade strains.