猪传染性胃肠炎病毒qRT-PCR检测方法的建立  

Establishment of qRT-PCR for detection of transmissible gastroenteritis virus

作　　者：原冬伟[1] 颜子涵 申贵男 王一 李明月[1] YUAN Dong-wei;YAN Zi-han;SHEN Gui-nan;WANG Yi;LI Ming-yue(College of Animal Sci-ence and Veterinary Medicine,Heilongjiang Bayi Agricultural University,Daqing,Hei-longjiang 163319,China)

机构地区：[1]黑龙江八一农垦大学动物科技学院,黑龙江大庆163319

出　　处：《中国兽医学报》2020年第10期1913-1917,共5页Chinese Journal of Veterinary Science

基　　金：黑龙江八一农垦大学引进人才科研启动计划资助项目(XYB2014-11);黑龙江八一农垦大学三横三纵支持计划资助项目(ZRCPY201907);黑龙江省自然科学基金资助项目(C2018-049,QC2015121);黑龙江省大学生创新创业训练计划资助项目(201810223047)。

摘　　要：猪传染性胃肠炎(TGE)是由猪传染性胃肠炎病毒(TGEV)感染引起猪以腹泻、呕吐、脱水为主要特征的一种高度接触性肠道传染病,可造成极高的仔猪死亡率,且临床剖检很难与其他病毒性腹泻区别。本试验参考TGEV JS2012株N基因保守序列设计引物,构建阳性标准品,建立qRT-PCR检测方法。将阳性标准品10倍倍比稀释构建标准曲线,结果显示,在6.06×10^2~6.06×10^8拷贝/μL扩增具有良好的线性关系。同时对临床生产中常见的腹泻病毒:轮状病毒、猪德尔塔冠状病毒、猪流行性腹泻病毒核酸进行扩增,结果均为阴性,无交叉反应,表明该方法具有良好的特异性。通过与传统PCR检测方法对比,本试验建立的检测方法具有更高的灵敏性,为临床检测TGEV的早期感染和预防提供技术支持。Transmissible gastroenteritis(TGE) is a highly contagious intestinal infectious disease(transmissible gastroenteritis virus) that causes diarrhea,vomiting and dehydration in pigs,which caused highly mortality in piglets,and difficult to distinguish from other viral diarrhea diseases.In this study,primers were designed according to the conserved sequence of N gene of TGEV JS2012 strain,and the positive standard was constructed to establish qRT-PCR method.The standard curve was constructed by 10-fold specific dilution of the positive standard,and the results showed that it has a good linear relationship in the range of 6.06×10^2 copies/μL and 6.06×10^8 copies/μL.At the same time,the common diarrhea viruses in clinical production,such as rotavirus,porcine delta coronavirus and porcine epidemic diarrhea virus,were detected as negative and there was no cross reaction,which indicated that the method had a good specificity.Compared with the conventional PCR method in the laboratory at present,the detection method established in this study has higher sensitivity.The establishment of this method provides a technical support for clinical detection of early infection and prevention of TGEV.

关 键 词：QRT-PCR 猪传染性胃肠炎病毒 N基因 

分 类 号：S852.65[农业科学—基础兽医学]