Abstract
目的 对单管-套式多重聚合酶链反应(PCR)鉴定间日疟原虫基因型方法的应用价值进行研究.方法以间日疟原虫环子孢子蛋白(CSP)基因为模型,综合套式多重PCR、等位特异PCR和标签引物扩增等技术,使用Primer Premier 5.0软件和NCBI的BLAST网络改进与优化引物设计,并用矩阵试验法优化反应条件和程序,同时,用单管-套式多重PCR和套式PCR对71份间日疟患者血标本的间日疟原虫基因型鉴定结果作比较. 结果单管-套式多重PCR可同时扩增出3种不同长度的基因型/族特异片段,且基本消除了二聚体等噪音.用单管-套式多重PCR鉴定检测71份间日疟患者血标本,62份与套式PCR鉴定结果一致,并新发现6份不同基因型和PV-Ⅱ型的混合感染;还将2份套式PCR鉴定为热带族的血标本纠正为PV-Ⅱ型,1份阴性血样鉴定为温带族.结论研究建立的单管-套式/多重PCR鉴定间日疟原虫基因型的方法,简化了试验步骤,节省了试剂耗材,消除了PCR噪音,提高了检出间日疟原虫混合感染和低度感染的敏感性;是一种具有发展潜力的检测单核苷酸多态性和基因型鉴定的方法。
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