生物素标记寡核苷酸探针检测和鉴定单核细胞增多症李氏菌的研究

Biotin-labeled Oligonucleotide Probe for Detection and Identification of L. monocytogenes

采用寡核苷酸生物素探针进行了斑点杂交试验。结果表明,12株单核细胞增多症李氏菌均与探针呈杂交阳性,其它种李氏菌和细菌无一与探针杂交。探针检测靶DNA的敏感性为10ng,过夜增菌培养,可提高敏感性10~3～10~4倍,测出样品中少至150个细菌,探针法检测市售猪肉和牛奶的阳性率高于分离培养法,具有推广应用价值。

A novel and practical dot blot hybridization assay was developed for detection and identification of L. monocytogenes . The biotin-labeled synthetic oligonucleotide probe was considered specific for the listeriolysin gene of L. monocytogenes . Using the probe, we were able to detect as little as 10 ng of DNA and as few as 150 bacterial cells in the simulated samples cultured overnight. It reacted with all 12 strains of L. monocytogenes tested, and no cross-reaction with other species of Listeria and bacteria took place. Dot blot hybridization got more positive results than bacterial cultures in detection of L. monocytogenes in the pork and milk samples obtained from the market. It is concluded that this method is applicable to detect L. monocytogenes in animal food.