结肠弯曲菌Real-time PCR检测方法的建立  

Real-time PCR development for identification of Campylobacter coli from stool specimen

作　　者：乔博[1] 许学斌[1] 顾一心[1] 刘国栋[1] 赵飞[1] 何利华[1] 张建中[1] 张茂俊[1] 

机构地区：[1]中国疾病预防控制中心传染病所,国家传染病重点实验室,北京102206

出　　处：《中国人兽共患病学报》2012年第10期969-972,共4页Chinese Journal of Zoonoses

基　　金：Supported by the National Natural Science Foundation（No.81071314）~~

摘　　要：目的建立结肠弯曲菌实时PCR检测方法并评价该方法的可行性。方法BLAST与Vector NTI Suite 6.0筛选出结肠弯曲菌的特异基因ceuE作为检测靶基因,Primer 5.0和Oligo6.0设计引物和探针,优化实时PCR反应体系并评价其最低检测限、特异性、可重复性。结果与结论在结肠弯曲菌的ceuE特异基因上设计引物和探针,经优化了的实时PCR反应体系最低检测限为5.62CFU,特异度为100%,变异系数为0.15%～1.30%,本研究为结肠弯曲菌的快速检测提供了一种参考方法。The aim of this study is to develop the real-time PCR assays to detect and qualify C.coli from stool specimen.The primers and probe were designed based on the specific sequence of ceuE gene in C.coli using Primer 5.0 and Vector NTI Suite 6.0 e.The PCR assay was optimized with the reference C.coli strains.The standard curve based on the dilutions of genomic DNA showed a linear relationship between log CFU/mL and threshold cycles.The detectable limitation was 5.62 CFU/mL by using purified DNA from bacteria culture.The reproducibility of this assay was assessed by calculating the variation of the threshold cycle value and the slope from test repeats for the same strains and different strains.Our results indicate that the developed assay has high sensitivity and specificity for identification of C.coli.

关 键 词：结肠弯曲菌 实时PCR ceuE 

分 类 号：R378[医药卫生—病原生物学]