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
袁琳凯 1马崇欢 1李丁山 1陈志炜 1江宵烽 1丁新伦 1张洁 1吴祖建1
作者信息
- 1. 福建农林大学植物病毒研究所/闽台作物有害生物生态防控国家重点实验室,福建 福州 350002
- 折叠
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.
关键词
柑橘黄化脉明病毒/柑橘衰退病毒/啤酒花矮化类病毒/多重RT-PCRKey words
Citrus yellow vein clearing virus/citrus tristeza virus/hop stunt viroid/multiplex RT-PCR引用本文复制引用
基金项目
福建省三明市横向项目(KH210264A)
福建省三明市横向项目(KH210295A)
福建省三明市横向项目(KH220062A)
出版年
2024
国家科技期刊平台
NSTL
万方数据