环介导等温扩增技术快速检测犬细小病毒方法的建立  

Rapid Diagnosis of Canine Parvovirus Infection by Loop-Mediated Isothermal Amplification Method

作　　者：温海 秦海斌 朱骞 贺星亮 陆承平[2] 张汇东 

机构地区：[1]公安部南京警犬研究所,江苏南京210012 [2]南京农业大学预防兽医学重点实验室,江苏南京210095

出　　处：《动物医学进展》2012年第2期9-13,共5页Progress In Veterinary Medicine

基　　金：公安部应用创新计划(2007YYCXNJJQS161)

摘　　要：旨在建立一种利用环介导等温扩增技术(LAMP)检测犬细小病毒感染的新方法。根据Gen-Bank中犬细小病毒(CPV)VP2基因序列,设计4条LAMP特异性引物,对反应条件、特异性、敏感性、可视化效果和应用效果进行研究。结果显示,在65℃等温条件下、1h内可完成LAMP扩增过程;病毒的最低检出限量为10-2个TCID50/mL;特异性和可视效果良好;对36份临床标本进行检测,阳性检出率为80.5%(29/36),检出率高于普通PCR 72.2%(26/36)。建立的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,为临床检测犬细小病毒感染提供了一种快速简便的新方法。A newly method for rapid diagnosis of canine parvovirus infection was developed by loop-mediated isothermal amplification method.According to the published GenBank sequence,four strips of primers specific recognising vp2 gene of canine parvovirus were designed.We studied the optimal reaction conditions and estemblished LAMP method for detecting CPV viruses from feces.Then we examined it′s specificity,sensitivity,the visual effect and efficient in field detection.The amplication can be performed at a constant temperature 65 ℃ within 1 hour.The detection limit was 10-2 TCID50/mL CPV.The results of LAMP could be visualized exactly by nake eyes after stained by SYBR Green Ⅰ.In 36 fecal samples which were selected from dogs suspectly infected with canine parovirus,29 samples（80.5%）were detected positive.This test demonstrated the high superiority on specificity,sensitivity,efficiency and convenience of LAMP method.Compared with conventional PCR method,LAMP had a higher detective proportion and didn＇t need complex facilities during performing.Therefore,this paper provides a newly excellent rapid method for the diagnosis of canine parvovirus infection.

关 键 词：环介导等温扩增 犬细小病毒 快速检测 

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