目的建立快速、灵敏地检测奶粉中沙门氏菌的TaqMan探针实时荧光PCR方法。方法根据沙门氏菌的inv A基因序列设计引物与TaqMan探针,建立并评价实时荧光PCR反应体系的特异性,制备鼠伤寒沙门菌标准株(CMCC 50115)10倍梯度稀释的菌液,检测对纯菌液的灵敏度,制备染菌量分别为1.4×(100~105)CFU/25 g奶粉的样本进行人工染菌量试验,在增菌0、4、8、12、16、20、24、36和48 h时各取1 m L培养液,提取DNA进行TaqMan实时荧光PCR检测,同步进行传统细菌分离,比较2种方法对人工染菌标本的最低检测限。结果建立的沙门氏菌Taq Man探针实时荧光PCR仅需40min即可完成检测,与志贺氏菌、大肠埃希氏菌等非目标细菌无交叉反应,对纯菌检测下限为1.4 CFU/m L,对染菌量为1.4×100CFU/25 g奶粉的检测下限仅需增菌16h即可检测出来,而要达到这个检测限,传统方法则需要36h。结论沙门氏菌TaqMan探针实时荧光PCR快速、特异、灵敏,可用于婴幼儿奶粉中沙门氏菌的快速筛查,提高食品安全风险监测的速度和灵敏度。 Objective To establish a real-time fluorescent PCR method for rapid and sensitive detection of Salmonella in milk powder by TaqMan probe. Methods According to the sequence of inv A gene of Salmonella, primers and TaqMan probes were designed and used to establish and evaluate the specificity of real-time fluorescence PCR reaction system. The 10-fold dilutions of Salmonella typhimurium standard strain (CMCC 50115) Liquid sensitivity, the preparation of the amount of bacteria were 1.4 × (100 ~ 105) CFU / 25 g milk samples were tested for the amount of artificial bacteria in the enrichment 0,4,8,12,16,20,24,36 and At 48 h, 1 m L of each culture medium was taken and DNA was extracted for real-time PCR detection by TaqMan. Conventional bacterial isolation was performed simultaneously. The minimum detection limits of the two methods for artificial bacterial contamination were compared. Results The real-time fluorescent PCR of Salmonella TaqMan probe was established in 40 min, and no cross-reaction with non-target bacteria such as Shigella and Escherichia coli was detected. The detection limit was 1.4 CFU / m L, The detection limit of 1.4 × 100CFU / 25 g milk powder could be detected only by adding bacteria for 16h. To reach this detection limit, the traditional method requires 36h. Conclusion The real-time fluorescent PCR of Salmonella TaqMan probe is rapid, specific and sensitive. It can be used for rapid screening of Salmonella in infant formula and improve the speed and sensitivity of food safety risk monitoring.