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当前位置: 首页 > 文章 > 猪乙型脑炎病毒Ⅰ、Ⅲ型的RT-LAMP检测方法建立与应用 畜牧与兽医 2015,47 (5) 26-30
Position: Home > Articles > Establishment and application of LAMP for swine Japanese encephalitis virus type Ⅰ and Ⅲ Animal Husbandry & Veterinary Medicine 2015,47 (5) 26-30

猪乙型脑炎病毒Ⅰ、Ⅲ型的RT-LAMP检测方法建立与应用

作  者：
李海洋;殷秀辰;饶柏钟;刘惠莉
单  位：
上海农业科学院畜牧兽医研究所
关键词：
猪;乙型脑炎病毒;环介导等温扩增;诊断
摘  要：
参考Gen Bank中猪乙型脑炎病毒(JEV)Ⅰ型和Ⅲ型毒株的E基因序列,通过比对分析,选取8个保守区域设计了6条引物,经条件优化及临床样品验证,本研究建立了能同时检测JEVⅠ型和Ⅲ型毒株E基因的RT-LAMP方法。该方法的最低检测灵敏限度为15 copies,具有较好的特异性,对猪常见病原如猪流感病毒(SIV)、猪瘟病毒(CSFV)、猪传染性胃肠炎病毒(TGEV),猪繁殖与呼吸综合征病毒(PRRSV),猪细小病毒(PPV)和伪狂犬病病毒(PRV)等均没有特异性扩增。与RT-PCR检测方法相比,其灵敏性高10倍。用本研究建立的RT-LAMP方法检测来自屠宰场的1 500份猪血清,阳性检出率为0.4%,与RT-PCR吻合率达100%,表明本试验建立的RT-LAMP方法适用于Ⅰ、Ⅲ基因型JEV的快速、批量筛检,为JEV感染的流行病学调查提供了必要的技术手段。
译  名：
Establishment and application of LAMP for swine Japanese encephalitis virus type Ⅰ and Ⅲ
作  者：
LI Haiyang;YIN Xiuchen;RAO Baizhong;LIU Huili;College of Fisheries and Life Science,Shanghai Ocean University;Institute of Animal Science and Veterinary Medicine,Shanghai Academy of Agricultural Sciences;Key Laboratory of Agricultural Genetics and Breeding;
关键词：
swine;;Japanese encephalitis virus;;loop-mediated isothermal amplification;;diagnosis
摘  要：
Six primers were designed to spot eight specific regions of a target gene according to the sequences of Japanese encephalitis virus(JEV) E gene published in Gen Bank. A RT- LAMP detection method for JEV genotypes Ⅰ and Ⅲ was established by optimizing the reaction conditions. The sensitivity limit of RT- LAMP was 15 copies. The method showed specific amplification for JEV,while the common pathogens such as swine flu virus( SIV),classical swine fever virus( CSFV),porcine transmissible gastroenteritis virus( TGEV),porcine respiratory and reproductive syndrome virus( PRRSV),swine parvovirus( PPV) and porcine pseudorabies virus( PRV) could not be detected. Compared to RT- PCR,the coincidence rate was 100% and the sensitivity was ten- fold higher in agarose gel electrophoresis. RT-LAMP established in this study was used to detect 1 500 serum samples of pigs from the slaughter- house. The positive rate was 0. 4%,which was 100% in agreement with that of RT- PCR,indicating that the established LAMP was suitable for clinical detection.

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