CAJ | 학술논문

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.
목적 건립결장만곡균실시PCR검측방법병평개해방법적가행성.방법 BLAST여Vector NTI Suite 6.0사선출결장만곡균적특이기인ceuE작위검측파기인,Primer 5.0화Oligo6.0설계인물화탐침,우화실시 PCR반응체계병평개기최저검측한、특이성、가중복성.결과 여결론 재결장만곡균적ceuE특이기인상설계인물화탐침,경우화료적실시PCR반응체계최저검측한위5.62CFU,특이도위100%,변이계수위0.15%~1.30%,본연구위결장만곡균적쾌속검측제공료일충삼고방법.
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.