• 中文核心期刊
  • CSCD来源期刊
  • 中国科技核心期刊
  • CA、CABI、ZR收录期刊

柑橘3种病毒类病原多重RT-PCR检测技术的建立及应用

A Multiplex RT-PCR Assay for Detecting Three Pathogens Infecting Citrus Plants

    摘要
  • 摘要:
      目的  建立柑橘黄化脉明病毒（citrus yellow vein clearing virus, CYVCV）、柑橘衰退病毒（citrus tristeza virus, CTV）和啤酒花矮化类病毒（hop stunt viroid, HSVd）的多重RT-PCR检测体系。
      方法  设计多重RT-PCR引物，分析其特异性，确定其最佳浓度比、最适退火温度及灵敏度，在此基础上对福建地区的柑橘样品进行检测。
      结果  确定了CYVCV-F/R、CTV-F/R和HSVd-F/R等3对引物的最佳浓度比例为1∶1∶2，最适退火温度为52.9 ℃，灵敏度结果显示该体系可检测模板稀释到10−2的阳性样品。应用该体系对采自福建部分地区的157份柑橘样品进行检测，结果发现，CYVCV、CTV和HSVd的检出率分别为47.1%、56.7%和22.9%。
      结论  成功建立了柑橘CYVCV、CTV和HSVd病原的多重RT-PCR检测方法，为该类病害的检测提供准确、快速的检测方法。

     

    Abstract:
      Objective   A multiplex RT-PCR assay to simultaneously detect citrus yellow vein clearing virus (CYVCV), citrus tristeza virus (CTV), and hop stunt viroid (HSVd) was developed.
      Method   Multiplex RT-PCR primers were designed, and their specificity was analyzed. The optimal concentration ratio, annealing temperature and sensitivity of primers were determined. Moreover, citrus samples from Fujian region were detected by the established multiplex RT-PCR.
      Result   The optimized concentration ratio of CYVCV-F/R, CTV-F/R, and HSVd-F/R primers was 1:1:2, and the annealing temperature at 52.9 ℃ for the assay. The assay displayed a sensitivity at 10-2 dilution and the positive detections on CYVCV of 47.1%, on CTV of 56.7%, and on HSVd of 22.9% on 157 detected samples.
      Conclusion  A rapid, accurate, applicable multiplex RT-PCR method for CYVCV, CTV, and HSVd detections was successfully established.

     

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