基于ORF7基因的猪繁殖与呼吸综合征病毒RT-RAA检测方法的建立  

Establishment of RT-RAA Method for Detection of PRRSV Based on ORF7 Gene

作　　者：于娜 马佳镁 张紫薇 黄春媛 范悦轩 郑佳馨 张艳[1,3] 刘光亮 曹宗喜 YU Nal;MA Jia-meil;ZHANG Zi-wei;HUANG Chun-yuan;FAN Yue-xuanl;ZHENG Jia-xin;ZHANG Yan;LIU Guang-liang;CAO Zong-xi(Hainan Provincial Key Laboratory of Tropical Breeding and Epidemiological Diseases,Institute of Animal Husbandry and Veterinary Medicine,Hainan Academy of Agricultural Sciences,Haikou,5711o0,China;Tarim Animal Disease Diagnosis and Prevention Engineering Laboratory of Xinjiang Production and Construction Corps,College of Animal Science and Technology,Tarim University,Aral,Xinjiang,843300;College of Veterinary Medicine,Xinjiang Agricultural University,Urumchi 830000,China;Suzhou Mars Biotechnology Co,Ltd.,Suzhou,Jiangsu,215021,China)

机构地区：[1]海南省农业科学院畜牧兽医研究所、海南省热带动物繁育与疫病研究重点实验室,海南海口571100 [2]塔里木大学动物科学与技术学院、新疆生产建设兵团塔里木动物疫病诊断与防控工程实验室,新疆阿拉尔843300 [3]新疆农业大学动物医学学院,新疆乌鲁木齐830000 [4]苏州天隆生物科技有限公司,江苏苏州215021

出　　处：《动物医学进展》2024年第8期117-119,共3页Progress In Veterinary Medicine

基　　金：国家自然科学基金项目(32060796);海南省重点研发项目(ZDYF2023XDNY038);海南省重大科技计划项目(ZDKJ2021030);国家重点研发计划(2021YFA0805905)。

摘　　要：根据GenBank公布的ORF7基因保守序列设计了特异性引物,建立了PRRSVORF7基因反转录-重组酶介导等温扩增(RT-RAA)荧光法快速检测方法。结果表明,RT-RAA荧光法在42℃、20min内即可检测到PRRSV特异性扩增曲线,最低检测浓度为9.9×10^(6)copies/μL;与PRVBartha-K61、PCV、CSFV、PRRSVGD、PRRSVXH-GD、PRRSVGM2、PRRSVCH-1a病毒均无交叉反应。分别对157份样品进行荧光RT-RAA法与PCR方法的检测进行比对,结果显示,荧光RT-RAA法敏感性为94.0%,特异性为100%。建立的PRRSV荧光RT-RAA法操作简便,特异性强,灵敏度高。Specific primers were designed according to the conserved sequence of ORF7 gene published in GenBank,and a rapid detection method for reverse transcription recombinase mediated isothermal amplification(RT-RAA)fluorescence of PRRSV ORF7 gene was established.The specific amplification curve of PRRSV could be detected by RT-RAA fluorescence method within 20 min at 42℃.The lowest detection limit was 9.9×10^(6)copies/μL;There was no cross-reactivity with PRV Bartha-K61,PCV,CSFV,PRRSV GD,PRRSV XH-GD,PRRSV GM2,PRRSV CH-la strains.The fluorescence RT-RAA method and PCR method were compared with 157 samples,and the results showed that the fluorescence RT-RAA method had a sensitivity of 94.0%and a specificity of 100%.The PRRSV fluorescence RT-RAA method established in this study is easy to operate,has strong specificity and high sensitivity,and can quickly and effectively detect clinical samples.

关 键 词：猪繁殖与呼吸综合征病毒 ORF7基因 反转录-重组酶介导等温扩增 特异性 灵敏度 

分 类 号：S852.659.6[农业科学—基础兽医学]