PCR和实时荧光PCR方法检测华支睾吸虫

Detection of Clonorchis sinensis by PCR and Real-Time PCR

目的利用PCR技术建立快速、灵敏、特异的检测华支睾吸虫的方法,比较PCR和实时荧光PCR检测华支睾吸虫的灵敏性和特异性,探讨其在华支睾吸虫检测中的应用。方法根据华支睾吸虫的特异性基因序列,设计了PCR引物及实时荧光PCR的特异性引物和探针,分别进行灵敏性和特异性试验。结果设计的引物能够扩增华支睾吸虫,而且探针的特异性很高。实时荧光PCR对华支睾吸虫的最低检测浓度为3 fg/μl,实时荧光PCR的灵敏性比常规PCR高2个数量级。结论PCR和实时荧光PCR检测华支睾吸虫的准确性和特异性均高,操作简便,为华支睾吸虫感染的诊治和预防提供有效的技术手段和方法依据。

Objective To establish a quick,sensitive and special method for detecting and evaluating the accuracy and specificity of PCR and real-time PCR in detection of Clonorchis sinensis. Method According to the specific sequence of Clortorchis sinensis genes, the primers of PCR and real-time PCR were designed separately and the fluorogenic probe was designed and synthesized. Result The designed primers could amplified the DNA,and the probes had high specificity and stability. The detection limit of the real-time PCR was 3fg/ul, and the sensitivity of real-time PCR was higher than that of PCR. Conclusion The PCR and real-time PCR are rapid, simple methods with high sensitivity and specificity, and they provide an efficient way for diagnosis,treatment and control of Clonorchis sinensis infections.