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当前位置: 首页 > 文章 > 鸡源产气荚膜梭菌重组酶聚合酶扩增检测方法的建立 中国兽医科学 2019 (12) 1475-1483
Position: Home > Articles > Recombinase polymerase amplification:a detection method for the detection of Clostridium perfringens from chicken Chinese Veterinary Science 2019 (12) 1475-1483

鸡源产气荚膜梭菌重组酶聚合酶扩增检测方法的建立

作  者：
许笑;王艳华;MUmar Zafar Khan;王恒;蔡建平
单  位：
中国农业科学院兰州兽医研究所家畜疫病病原生物学国家重点实验室;江苏省动物重要疫病与人兽共患病防控协同创新中心
关键词：
产气荚膜梭菌;重组酶聚合酶扩增技术;荧光定量检测;鸡坏死性肠炎
摘  要：
为建立鸡源产气荚膜梭菌(Clostridium perfringens)的快速诊断方法,本研究以产气荚膜梭菌α-毒素的编码基因plc为基础设计、合成引物和探针,并以real-time PCR的方法进行验证,建立了鸡源产气荚膜梭菌的重组酶聚合酶扩增(RPA)检测方法。该方法可在38℃、25 min内对产气荚膜梭菌的α-毒素编码序列进行快速特异性扩增,最低检测限量可达1×10~1copies/μL,灵敏度良好。以此方法检测实验室保存的临床样本,其检测结果与real-time PCR的检测结果一致。上述结果表明,本研究建立的real-time RPA方法在检测鸡源产气荚膜梭菌时具有操作简单、快速灵敏的特点,适用于鸡源产气荚膜梭菌的临床检测。
译  名：
Recombinase polymerase amplification:a detection method for the detection of Clostridium perfringens from chicken
作  者：
XU Xiao;WANG Yan-hua;M Umar Zafar Khan;WANG Heng;CAI Jian-ping;State Key Laboratory of Veterinary Etiological Biology/Lanzhou Veterinary Research Institute,Chinese Academy of Agricultural Sciences;JiangSu Co-Innovation Center for Prevention and Control of Animal Infectious Disease and Zoonoses;
关键词：
Clostridium perfringens;;recombinase polymerase amplification;;real-time PCR;;chicken;;necrotic enteritis
摘  要：
In order to develop a rapid molecular detection method for Clostridium perfringens(C.perfringens) causing chicken necrotic enteritis(NE),a real-time recombinase polymerase amplification(RPA) method based on the α-toxin gene of C.perfringens(Cp PLC) was built,and validated with real-time PCR method as a control.This real-time RPA can complete the specific amplification to Cp PLC at 38 ℃ for 25 min, with lower reaction temperature and shorter run time compared to a common real-time PCR method,as well as high sensitivity in lowest detectable copy number to 1×10~1 copies/μL.It was also found that the results for clinical sample detections with this real-time RPA are highly consistent with the conventional real-time PCR.Our results shows that the real-time RPA detection for C.perfringens established here can be used for detecting C.perfringens related to chicken NE.

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