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根据基因库中单孢子虫的基因保守序列,设计了1对特异性引物和1条TaqMan探针,对反应条件和试剂浓度进行优化,建立了检测单孢子虫的荧光定量PCR方法。该方法对单孢子虫的检测敏感性达到40拷贝数,比常规PCR敏感性高100倍;对派琴虫、折光马尔太虫、荧光假单胞菌、副溶血弧菌、溶藻弧菌、河弧菌和拟态弧菌等病原体的检测结果全为阴性。表明该研究建立的单孢子虫荧光定量PCR具有特异、敏感、快速、定量和重复性好等优点,可用于临床单孢子虫感染的快速检测。
According to the conserved sequences of single sporozoites in the gene bank, one pair of specific primers and one TaqMan probe were designed to optimize the reaction conditions and reagent concentration, and a fluorescence quantitative PCR method was established to detect the sporozoites. The method has a sensitivity of 40 copies to the detection of the sporozoites, which is 100 times higher than that of the conventional PCR; Vibrio River and mimic Vibrio and other pathogens test results were all negative. The results showed that the fluorescence quantitative PCR of the sporozoites established in this study is specific, sensitive, rapid, quantitative and reproducible. It can be used for the rapid detection of clinical infection of sporozoites.