欢迎访问《中国兽医科学》网站 -- www.zgsykx.com　　　　　　　　　　

　 　 　


猪生殖与呼吸综合征病毒TaqMan荧光定量RTPCR
检测方法的建立

宋志军1，2， 宋长绪2， 杨增岐1， 朱道中3， 武占银4， 
蒋智勇2， 林志雄3， 刘燕玲2， 张福良1，2

(1. 西北农林科技大学 动物科技学院， 陕西 杨凌712100； 2. 广东省农业科学院 兽医研究所， 广东 广州
510640； 3.广州出入境检验检疫局， 广东 广州510623； 4.宁夏动物防疫站， 宁夏 银川750002)

摘要：  
根据GenBank登录的PRRSV保守基因序列设计合成了引物和探针,并对其进行了筛选;对荧光定量PCR的反应条件进行了优化,建立了TaqMan荧光定量RTPCR检测方法。同时用建立的检测方法对组织病料进行了检测,并与常规RTPCR做了对比。结果显示，所建立的TaqMan荧光定量RTPCR方法灵敏度可达50×100拷贝/μL,比常规RTPCR灵敏度高100倍。用该方法对东莞、增城、湛江等地的猪血清和多种组织样品进行了检测。结果，该方法与常规RTPCR检测方法的阳性符合率为100%。用该方法对3份不同的组织样品进行了重复检测,结果表明，该方法具有良好的重复性,可满足当前PRRS的诊断需要。
关键词：  
猪生殖与呼吸综合征病毒; 实时定量RTPCR; TaqMan荧光探针
中图分类号：S 852.659.6: Q 503  文献标识码： A  文章编号：1673-4696(2006)02-0098-05

Development of realtime TaqManquantitative RTPCR assay for 
detection of porcine reproductive and respiratory syndrome virus

SONG Zhijun1.2, SONG Changxu2, YANG Zengqi1, ZHU Daozhong3, WU Zhanyin4, 
JIANG Zhiyong2, LIN Zhixiong3, LIU Yanling2, ZHANG Fuliang1,2

(1. College of Animal Science and Technology, Northwest SciTech University of Agriculture and Forestry,
Yangling 712100,China; 2. Research Institute of Veterinary Medicine, Guangdong Academy of Agricultural
Sciences, Guangzhou 510640,China; 3. Guangzhou EntryExit Inspection and Quarantine Bureau, 
Guangzhou 510623,China; 4. Ningxia Animal Quarantine Station, Yinchuan 750002,China)
Abstract: The probes and primers were designed and synthesized according to the conserved gene ORF7 of PRRSV available in GenBank, and then reaction parameters were optimized to develop a realtime TaqManquantitative RTPCR assay. The serum and other tissue samples from pigs on farms in Dongguan, Zengcheng， Zhanjiang and other areas of Guangdong Province were detected by using the established quantitative RTPCR assay, and the results was compared with that of routine RTPCR. The developed quantitative RTPCR assay could detect 50×100 copy/μL of plasmid DNA and its sensitivity was 100 times higher than that of the routine RTPCR, while the results of the quantitative RTPCR were the same as that of the routine RTPCR. Three samples were examined using the realtime RTPCR repeatedly and the results indicated that the realtime quantitative RTPCR was reproducible and could be used for the diagnosis of PRRSV infection.

Key words: PRRSV; realtime quantitative RTPCR; TaqMan fluorescence probe

《中国兽医科学》 © 版权所有
Chinese Veterinary Science
zgsykx@zgsykx.com