作 者：
保雨;聂福平;王昱;杨俊;袁曾壮;叶自霞;刘亚娟;张姜琳;吴蕊
单 位：
西南大学动物科技学院;重庆出入境检验检疫局重庆市进出口食品安全工程中心
关键词：
流产嗜衣原体;TaqMan-MGB;荧光定量PCR
摘 要：
为建立一种快速、准确检测流产嗜衣原体(C.abortus)Taq Man-MGB荧光定量PCR方法,本研究根据C.abortus主要外膜蛋白基因的特异保守序列设计引物及探针,并优化反应条件,建立了检测C.abortus的荧光定量PCR方法。结果表明,以重组质粒为标准品建立的标准曲线在1.6×103拷贝/μL~1.6×107拷贝/μL内具有良好的线性关系,相关系数为0.9999。该方法仅对C.abortus的靶基因扩增呈阳性,而对鹦鹉热嗜衣原体、家畜嗜衣原体、鼠衣原体、沙眼衣原体、肺炎嗜衣原体、猪源衣原体核酸扩增结果均为阴性,特异性强;其最低检出限为1.6拷贝/μL;组内和组间重复性试验变异系数均小于3%,具有良好的重复性。利用建立的方法和普通PCR方法同时对225份临床样品进行检测,结果显示荧光定量PCR检出率比普通PCR高4.5%,表现较高的灵敏度和准确性。本研究建立的方法对C.abortus的临床鉴别检测和疾病诊断具有重要意义。
译 名：
Establishment of TaqMan-MGB real-time PCR for detecting Chlamydophila abortus
作 者：
BAO Yu;NIE Fu-ping;WANG Yu;YANG Jun;YUAN Zeng-zhuang;YE Zi-xia;LIU Ya-juan;ZHANG Jiang-lin;WU Rui;TIAN Jun;WANG Guo-ming;LIU Li;LI Ying-guo;College of Animal Science and Technology, Southwest University;Chongqing Import and Export Food Safety Engineering Center, Chongqing Entry-exit Inspection and Quarantine Bureau of China;College of Bioengineering, Chongqing University;College of Pharmacy and Bioengineering, Chongqing University of Technology;
关键词：
Chlamydophila abortus;;Taq Man-MGB;;real-time PCR
摘 要：
In this study, a Taq Man-based real time PCR assay was established for detecting Chlamydophila abortus with primers and Taq Man probe targeting the major outer membrane protein(MOMP) gene of C.abortus. Under the optimum condition,the results showed that the standard curve established with positive plasmid had a liner response from 1.6 ×103copies/μL to 1.6 ×107copies/μL with the correlation coefficient of 0.9999. The assay was specific and sensitive for detecting C.abortus with the detection limit of 1.6 copies/μL, but no amplification for C.psittaci, C.pecorum, C.trachomatis, C.pneumoniae, C.suis and C.muridarum. The intra- and inter-assay indicated that the method was repeatable and stable with the coefficients of variation less than3%. The established assay and conventional PCR were used to detect C.abortus in 225 clinical samples, and the detection rate of the real-time PCR was higher than conventional PCR. The method established in present study would be useful for experimental research and detection of C.abortus in clinical samples.
