[1]华明艳,宋兰芳,付成龙,等.基于微滴式数字PCR技术检测番茄褐色皱纹果病毒方法的建立与评价[J].江苏农业学报,2026,42(05):929-939.[doi:doi:10.3969/j.issn.1000-4440.2026.05.007]
　HUA Mingyan,SONG Lanfang,FU Chenglong,et al.Establishment and evaluation of a method for detecting tomato brown rugose fruit virus based on droplet digital PCR technology[J].,2026,42(05):929-939.[doi:doi:10.3969/j.issn.1000-4440.2026.05.007]
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基于微滴式数字PCR技术检测番茄褐色皱纹果病毒方法的建立与评价()

江苏农业学报[ISSN:1006-6977/CN:61-1281/TN]

卷:
42
期数:
2026年05期
页码:
929-939
栏目:
植物保护
出版日期:
2026-05-31

文章信息/Info

Title:
Establishment and evaluation of a method for detecting tomato brown rugose fruit virus based on droplet digital PCR technology
作者:
华明艳1; 宋兰芳1; 付成龙2; 崔少杰1; 孙海波1; 靳力争1; 金凤媚1
(1.天津市农业科学院现代都市农业研究所,天津300192；2.山东蒙恩现代农业发展有限公司,山东聊城252000)
Author(s):
HUA Mingyan1; SONG Lanfang1; FU Chenglong2; CUI Shaojie1; SUN Haibo1; JIN Lizheng1; JIN Fengmei1
(1.Institute of Modern Urban Agriculture, Tianjin Academy of Agricultural Sciences, Tianjin 300192, China；2.Shandong Meng’en Modern Agricultural Development Co.,Ltd., Liaocheng 252000, China)
关键词:
番茄褐色皱纹果病毒; 微滴式数字PCR; 检测方法; 灵敏度
Keywords:
tomato brown rugose fruit virus; droplet digital PCR; detection method; sensitivity
分类号:
S641.2
DOI:
doi:10.3969/j.issn.1000-4440.2026.05.007
文献标志码:
A
摘要:
番茄褐色皱纹果病毒(Tomato brown rugose fruit virus,ToBRFV)病作为威胁全球番茄产业的重要病害,具有早期无症状、传播速度快、危害程度重等特点。为建立ToBRFV的高效精准检测方法、实现番茄褐色皱纹果病毒病的有效防控,本研究通过设计 ToBRFV CP 基因保守区域特异性引物,构建质粒标准品,建立基于微滴式数字 PCR(ddPCR)的 ToBRFV 检测方法(ddPCR法),并将ddPCR法与实时荧光定量 PCR(qPCR)检测方法进行系统比较。结果表明,ddPCR法对ToBRFV的检测下限为1 μL 1.9×100拷贝,而qPCR方法对ToBRFV的检测下限为1 μL 1.9×102拷贝,且ddPCR法仅对 ToBRFV 呈现阳性扩增,而对番茄花叶病毒(ToMV)、烟草花叶病毒(TMV)、番茄斑萎病毒(TSWV)等田间常见病毒无交叉反应。基于ddPCR 方法对30份田间疑似感染ToBRFV的样本进行检测,阳性检出率达 93.3%,显著高于qPCR方法的 76.7%。综上,本研究建立的ddPCR检测方法在 ToBRFV 早期诊断、低载毒量样本检测及田间精准筛查中表现出检测下限低、特异性强和漏检风险低等优势,可有效应用于ToBRFV的流行监测,为ToBRFV的科学防控及番茄产业健康发展提供技术支撑。
Abstract:
Tomato brown rugose fruit virus (ToBRFV) disease is an important disease that threatens the global tomato industry. It has the characteristics of early asymptomatic, rapid transmission and severe damage. In order to establish an efficient and accurate detection method for ToBRGFV and realize the effective prevention and control of tomato brown rugose fruit virus disease, this study designed specific primers for the conserved region of ToBRFV CP gene and constructed plasmid standards. The ToBRFV detection method based on droplet digital PCR (ddPCR) was successfully established, and the ddPCR method was systematically compared with the real-time fluorescence quantitative PCR (qPCR) detection method. The results showed that the detection limit of ddPCR for ToBRFV was 1.9×100 copies per microliter, while the detection limit of qPCR for ToBRFV was 1.9×102copies per microliter. The ddPCR method only showed positive amplification for ToBRFV, but had no cross-reactions with tomato mosaic virus (ToMV), tobacco mosaic virus (TMV) and tomato spotted wilt virus (TSWV). Based on the ddPCR method, 30 samples suspected to be infected with ToBRFV in the field were tested, and the positive detection rate was 93.3%, which was significantly higher than 76.7% of the qPCR method. In summary, the ddPCR detection method established in this study showed the advantages of low detection limit, strong specificity and low risk of missed detection in the early diagnosis of ToBRFV, the detection of low-load samples and the accurate screening in the field. It can be effectively applied to the epidemic monitoring of ToBRFV and provide technical support for the scientific prevention and control of ToBRFV and the healthy development of tomato industry.

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备注/Memo

备注/Memo:
收稿日期：2025-10-14基金项目：国家重点研发计划项目(2023YFD1201503)；天津市重点研发计划项目(23YFZCSN00160)；2025年农业主导产业自主创新科研项目(2025ZYCX012)；泰山产业领军人才工程(TSCY20231269)作者简介：华明艳(1983-),女,辽宁建平人,硕士,助理研究员,主要从事番茄及草莓分子育种研究。(E-mail)26764891@qq.com通讯作者：金凤媚,(E-mail)Jinfm414@163.com
更新日期/Last Update: 2026-06-17