Molecular identification of lactic acid bacteria in Chinese rice wine using species-specific multiplex PCR

Chinese rice wine (CRW), a national unique and traditional food, has more than 4,000 years of history and is very popular in China. Lactic acid bacteria (LAB) play important roles in the fermentation of CRW because of its strong lactic acid production and high acid tolerance. It would be of practical interest to isolate the LAB species from CRW and to develop a simple and effective method for its accurate identification. In this study, 11 CRW-related LAB strains, frequently occurring in the fermentation of CRW, were isolated or collected and analyzed with the nested specifically amplified polymorphic DNA (nSAPD)-PCR technique. Eleven species-specific sequence characterized amplified region (SCAR) markers were generated from 11 polymorphic nSAPD bands. Three multiplex PCR assays were further established for the joint use of the 11 species-specific SCAR markers. Thus, a simple rapid molecular method was developed based on the application of nSAPD, SCAR, and multiplex PCR techniques, which allowed a rapid and simultaneous detection of 11 CRW-related LAB species. The simple rapid PCR-based molecular method can be used as a useful tool for the identification of LAB species in the traditionally fermented foods, especially in the rapid developing CRW industry in China.