欢迎访问《中国农学通报》，
中国农学通报

中国农学通报 ›› 2013, Vol. 29 ›› Issue (15): 37-41.doi: 10.11924/j.issn.1000-6850.2012-2980

所属专题： 马铃薯

• 农学 农业基础科学 • 上一篇    下一篇

马铃薯A病毒液相芯片快速检测方法的建立

刘洪义 张永江 刘忠梅 辛言言   

  • 收稿日期:2012-09-03 修回日期:2012-11-14 出版日期:2013-05-25 发布日期:2013-05-25
  • 基金资助:
    国家质检总局科技计划项目

Preliminary Development of Liquichip Detection Technique for Rapid Detection of Potato Virus A

  • Received:2012-09-03 Revised:2012-11-14 Online:2013-05-25 Published:2013-05-25

PDF (PC)

785

摘要： 旨在建立可检测马铃薯A病毒(Potato virus A，PVA)的液相芯片快速检测技术，用Primer Premier 5.0软件对GenBank中PVA核苷酸序列进行分析，设计其特异性探针并用生物素标记。探针偶联荧光编码微球后与PVA的PCR产物杂交反应，用液相芯片检测仪检测荧光信号。结果显示该法具有较好的特异性，不与侵染马铃薯的番茄黑环病毒(Tomato black ring virus，TBRV)、马铃薯纺锤块茎类病毒(Potato spindle tuber viroid，PSTVd)、马铃薯Y病毒(Potato virus Y，PVY)及番茄斑萎病毒(Tomato spotted wilt virus，TSWV)反应；检测灵敏度约为10 pg/μL总RNA。该方法可用于PVA的快速检测。

关键词: 再生体系, 再生体系

Abstract: To develop liquichip method for detecting Potato virus A (PVA), the nucleotide sequence of PVA was analyzed by using the software Primer Premier 5.0, and specific probe labeled with biotin was prepared and coupled with fluorescence-coded microspheres, which was used for hybridization reaction to RT-PCR products of PVA. Liquichip detection method for PVA was established by using Bio-Rad Liquichip to detect fluorescence signal in the reaction system. The specificity of this method was evaluated by applying the proposed method to detect four viruses infecting potato, TBRV, Potato spindle tuber viroid (PSTVd), Potato virus A (PVA) and Tomato spotted wilt virus (TSWV). The results demonstrated better specificity to PVA and no reaction with other three viruses. The sensitivity of the method was 1 pg/μL of total RNA, which was equal to the conventional RT-PCR gel electrophoresis method.The successful detection of PVA in potato samples suggested the feasibility of this procedure for routine testing