用TaqMan探针实时定量PCR法快速检测鼠疫耶尔森菌

目的建立一种快速、准确、特异的定量检测鼠疫耶尔森菌的方法。方法针对鼠疫耶尔森菌特异的染色体标识序列设计引物和TaqMan探针，进行实时定量聚合酶链反应(polymerase chain reaction，PCR)分析，优化反应体系，检测其灵敏度、特异性和重复性。结果以EV76灭活培养物为对象，检测灵敏度可达每反应体系0．08CfU；30株非鼠疫菌株的检测结果表明，对照菌株均为阴性，检测特异性良好；对同一样品进行17次重复检测，其循环阈值的标准偏差为0．35。结论TaqMan探针法能够灵敏、特异、稳定地对鼠疫耶尔森氏菌进行定量检测，为鼠疫监控、诊断提供有力手段。

Rapid detection of Yersinia pestis with TaqMan probe

Objective To establish a rapid and specific quantitative PCR assay for detecting of Yersinia pestis. Methods The primers and TaqMan probe targeted at signature sequence of chromosomal DNA were designed and applied to real time PCR assays. The reaction system was optimized, and the sensitivity, specificity and reproducibility were evaluated. Results The optimal reaction system was determined. The sensitivity of this assay was 0.08 CFU per reaction, and the detection results of 30 strains of non-Y. pestis showed the high specificity of the reaction. The standard deviation of seventeen-times detection of one sample was 0.35. Conclusions This assay was sensitive and specific for rapid identification and quantitative detection of Y. pestis. It will be an effective method for the surveillance and diagnosis of bubonic plague.