犬瘟热病毒RT-LAMP检测方法的建立和初步应用  

Detection of Canine Distemper Virus by Reverse Transcriptase Loop-Mediated Isothermal Amplification Method

作　　者：温海 秦海斌 贺星亮 朱骞 高一龙 张汇东 

机构地区：[1]公安部南京警犬研究所分子生物学实验室,南京210012

出　　处：《中国比较医学杂志》2012年第8期51-55,共5页Chinese Journal of Comparative Medicine

基　　金：公安部应用创新计划(2007YYCXNJJQS161)

摘　　要：目的利用环介导等温扩增(LAMP)技术建立一种检测犬瘟热病毒感染的新方法。方法根据GenBank中NP基因序列,设计4条LAMP特异性引物,对反应条件、特异性、可视化效果和应用效果进行研究。结果在60℃等温的条件下、1 h内可完成RT-LAMP扩增过程;特异性和可视效果良好;对63份临床标本进行检测,阳性检出率为71.4%(45/63),检出率高于RT-PCR的63.5%(40/63)。结论建立的RT-LAMP检测方法,显示了较高的特异性和敏感性,而且兼具高效、快捷、可视化的优势,为临床检测犬瘟热病毒感染提供了一种快速简便的新途径。Objective To develop a new method for rapid detection of canine distemper virus by reverse transcriptase loop-mediated isothermal amplification method （RT-LAMP）. Method According to the published GenBank sequence, four strips of primers specific recognizing NP gene of canine distemper virus were designed. We studied the optimal reaction conditions, specificity, sensitivity, and its visualized judging method of products. Then we used 63 clinical specimens to verify its efficiency in field detection. Results The amplication can be performed at a constant temperature 600（： within 1 hour, Specific primers did not recognize genes of other virus commonly infecting canine. The results of RT- LAMP could be easily observed by naked eyes after SYBR green I staining. In 63 specimens collected from Nanjing and Shangdong, 45 samples（71.4qo ）were positive, but only 40 samples （63.5 % ） were positive using RT-PCR. Conclusion The results of our study demonstrate a higher superiority of specificity, sensitivity, efficiency and convenience of the RT- LAMP method, and provide a new detection method for canine distemper virus.

关 键 词：环介导等温扩增 犬瘟热病毒 检测 

分 类 号：S852.655[农业科学—基础兽医学] S858.292[农业科学—兽医学]