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浙江大学学报（农业与生命科学版）
浙江大学学报（农业与生命科学版）  2023, Vol. 49 Issue (5): 696-707    DOI: 10.3785/j.issn.1008-9209.2023.05.121
作物重要细菌和病毒病害专题     
甜瓜瓜类蚜传黄化病毒侵染响应基因鉴定
杨思语1(),宫子惠1,胡仲远1,2,张明方1,2,杨景华1,2()
1.浙江大学农业与生物技术学院，浙江 杭州 310058
2.浙江大学海南研究院，海南 三亚 572025
Identification of genes in response to cucurbit aphid-borne yellows virus infection in melon
Siyu YANG1(),Zihui GONG1,Zhongyuan HU1,2,Mingfang ZHANG1,2,Jinghua YANG1,2()
1.College of Agriculture and Biotechnology, Zhejiang University, Hangzhou 310058, Zhejiang, China
2.Hainan Institute of Zhejiang University, Sanya 572025, Hainan, China
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摘要：

瓜类蚜传黄化病毒（cucurbit aphid-borne yellows virus, CABYV）是侵染甜瓜（Cucumis melo）等葫芦科作物的重要病毒之一，严重影响作物产量和品质。响应CABYV侵染的关键基因鉴定可为甜瓜抗病毒病育种提供靶基因。本研究用构建的CABYV侵染性克隆载体接种‘西州蜜’甜瓜，并对0 dpi（days post inoculation，感染后时间）、5 dpi、10 dpi、15 dpi、20 dpi时的甜瓜进行病情鉴定与转录组分析。结果表明：‘西州蜜’在20 dpi时表现出叶片明显褪绿、黄化和增厚等典型症状；基于转录组测序鉴定到响应CABYV侵染的差异表达基因（differentially expressed genes, DEGs）为1 654个，其中上调基因677个，下调基因977个。对这些响应基因进行基因本体（gene ontology, GO）与京都基因和基因组数据库（Kyoto Encyclopedia of Genes and Genomes, KEGG）富集分析，发现其主要富集在植物-病原互作、光合作用、淀粉和蔗糖代谢、乙醛酸盐和二羧酸盐代谢等途径和过程。差异基因共表达与互作分析发现，在CABYV侵染后，病原防御反应过程中的关键基因RIN4表达量呈下调趋势，推测其负调控甜瓜对CABYV的侵染响应。本研究结果为解析CABYV侵染甜瓜后基因响应的分子机制和甜瓜抗CABYV育种提供了重要依据。
关键词： 甜瓜;  瓜类蚜传黄化病毒;  转录组测序;  响应基因    
Abstract:

Cucurbit aphid-borne yellows virus (CABYV) is one of the most important viruses infecting cucurbits, such as melon (Cucumis melo), and severely affects the yield and quality of crops. Identification of CABYV-responsive genes can provide target genes for breeding of melon resistant to the viral disease. In this study, we used the CABYV infectious cloning vector to inoculate the melon XZM, and the disease identification and transcriptome analysis in the melon XZM after CABYV inoculation were performed at 0 dpi (days post inoculation), 5 dpi, 10 dpi, 15 dpi and 20 dpi. The results indicated that melon leaves showed typical disease symptoms of leaf chlorosis, yellowing and leaf thickening at 20 dpi. A total of 1 654 differentially expressed genes (DEGs) in response to CABYV infection were identified by transcriptome sequencing analysis, including 677 up-regulated genes and 977 down-regulated genes. Gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses showed that the responsive genes were mainly enriched in plant-pathogen interaction, photosynthesis, starch and sucrose metabolism, glyoxylate and dicarboxylate metabolism, etc., pathways and processes. Co-expression and interaction analysis of DEGs revealed that RIN4, a key gene in pathogen defense responses, may negatively regulate responses to CABYV infection in melon. This study demonstrates the possible molecular mechanism of responses to CABYV infection and provides a basis for breading melon against CABYV.
Key words: melon    cucurbit aphid-borne yellows virus    transcriptome sequencing    responsive genes
收稿日期: 2023-05-12 出版日期: 2023-10-25
CLC:  S652.1  
基金资助: 海南省重点研发计划项目(ZDYF2021XDNY166);国家西甜瓜产业技术体系项目(CARS-25-17)
通讯作者: 杨景华     E-mail: siyu.yang@zju.edu.cn;yangjinghua@zju.edu.cn
作者简介: 杨思语（https://orcid.org/0000-0002-6394-4530），E-mail：siyu.yang@zju.edu.cn
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杨思语
宫子惠
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引用本文:

杨思语,宫子惠,胡仲远,张明方,杨景华. 甜瓜瓜类蚜传黄化病毒侵染响应基因鉴定[J]. 浙江大学学报（农业与生命科学版）, 2023, 49(5): 696-707.

Siyu YANG,Zihui GONG,Zhongyuan HU,Mingfang ZHANG,Jinghua YANG. Identification of genes in response to cucurbit aphid-borne yellows virus infection in melon. Journal of Zhejiang University (Agriculture and Life Sciences), 2023, 49(5): 696-707.

链接本文:

https://www.zjujournals.com/agr/CN/10.3785/j.issn.1008-9209.2023.05.121    或    https://www.zjujournals.com/agr/CN/Y2023/V49/I5/696

图1  甜瓜‘西州蜜’接种pCB301-CABYV后在5个时间点的叶片病征A.叶片正面；B.叶片背面。
图2  20 dpi时甜瓜叶片中CABYV的 CP 基因扩增情况M：DL2000 DNA分子质量标志物。1～18：接种pCB301-CABYV的甜瓜发病叶片；19：阴性对照叶片。
图3  被侵染甜瓜叶片中CABYV含量的ELISA结果1～18：接种pCB301-CABYV的甜瓜发病叶片；19：阴性对照叶片。
图4  各转录组样本的相关性分析与主成分分析
图5  pCB301-CABYV侵染甜瓜后各组间的差异表达基因数
图6  0 dpi与20 dpi处理组差异表达基因GO功能注释（A）和KEGG富集分析（B）
  
图7  不同处理组间共有显著差异表达基因分析A.维恩图；B. GO功能注释；C. KEGG富集分析。
图8  共有显著差异表达基因植物-病原互作网络
图9  CABYV侵染后甜瓜 MELO3C021057 基因在5个时间点的相对表达量箱线图上不同大写字母表示在P＜0.01水平差异有统计学意义。
  
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