3种实验动物相关病原液相芯片检测方法的建立

Development of Liquichip Detection Technique for Three Pathogens in Laboratory Animals

为建立快速高通量检测实验动物质量相关布鲁菌、沙门菌、弓形虫3种病原体的方法,根据其序列设计引物及探针,探针经修饰后与荧光编码微球偶联,将偶联后的探针与PCR产物杂交反应,通过液相芯片检测仪(Luminex200)检测荧光信号,分析实验动物感染布鲁菌、沙门菌和弓形虫的情况。结果显示:初步建立了可同时检测布鲁菌、沙门菌和弓形虫的液相芯片检测方法,可特异地检测出3种目标病原的基因荧光信号,未检测出其他相关病原基因荧光信号;检测灵敏度达50拷贝/反应。本研究所建立的液相芯片检测方法可快速检测3种实验动物相关重要人兽共患病原体,对公共安全卫生保障、进出境实验动物检疫具有重要意义。

In order to develop a liquichip technique for detection of pathogens that infect laboratory animals,specific primers and their corresponding oligonucleotide probes for Brucella,Salmonella,Toxoplasma gondii were designed based on their conserved sequences.The labeled probes were coupled with fluorescent beads and used for hybridization reaction to PCR products.The detection technique was developed by using Luminex200 to screen fluorescence signals from the reaction system.The results showed that the developed method only detected Brucella,Salmonella,T.gondii but not other pathogens.The limit of detection was 50 copies of nucleic acids.The development of this rapid detection method provided an useful tool for public health and entry-exit laboratory animal quarantine.