›› 2010, Vol. 37 ›› Issue (4): 76-79.

• 生物技术 • 上一篇    下一篇

非洲猪瘟病毒实时荧光定量PCR检测技术的研究与评价

郭少平1,2,刘建2,吴绍强2   

  1. (1.内蒙古农业大学, 呼和浩特 010018; 2.中国检验检疫科学研究院, 北京 100029)
  • 收稿日期:1900-01-01 修回日期:1900-01-01 出版日期:2010-04-20 发布日期:2010-04-20
  • 通讯作者: 吴绍强

Research and Assessment of Real-time Quantitative PCR Assay for the Detection of African Swine Fever Virus

GUO Shao-ping1,2,LIU Jian2,WU Shao-qiang2   

  1. (1.Inner Mongolia Agricultural University, Hohhot 010018, China;2.Chinese Academy of Inspection and Quarantine, Beijing 100029, China)
  • Received:1900-01-01 Revised:1900-01-01 Online:2010-04-20 Published:2010-04-20
  • Contact: WU Shao-qiang

摘要: 根据非洲猪瘟病毒(African swine fever virus,ASFV)K205R基因序列设计合成引物及TaqMan探针,通过优化引物浓度、退火温度和Mg2+浓度,建立基于K205R基因的ASFV实时荧光定量PCR(real-time quantitative PCR,qPCR)检测方法。对在猪肝脏组织DNA中掺入重组质粒制备的模拟样品进行扩增来确定所建方法的实际检测效率,并将OIE推荐的基于p72的qPCR调整后作为评价该方法检测效果的对照。试验结果表明,基于K205R基因的检测方法在检测模拟样品时扩增效率要优于基于p72的方法,而且敏感性和特异性强,适用于非洲猪瘟的快速检测、监测。

关键词: 非洲猪瘟病毒; 实时荧光定量PCR; K205R; 检测; 研究; 评价

Abstract: A real-time quantitative PCR(qPCR)assay for the detection of African swine fever virus(ASFV)has been developed by designing the primers and TaqMan probe according to the sequence of the gene K205R. The reaction solutions and conditions were optimized by using different concentrations of the primers and Mg 2+ and different annealing temperature. The mask samples were adopted to verify its actual detective efficiency by adding the plasmid DNA into swine liver tissue. The qPCR detective method of ASFV based on p72 recommended by OIE was selected and modified to be as control method. Results showed that the amplification efficiency of the developed qPCR based on detection of K205R was better than that based on p72 and was sensitive and specific enough to detect African swine fever virus infection.

Key words: African swine fever virus; real-time quantitative PCR; K205R; detection; research; assessment

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