沃柑叶片响应柑橘溃疡病菌侵染的转录组分析  

Transcriptome analysis of Orah leaves in response to citrus canker infection

作　　者：邱发发 潘贞珍 李鸾翔 夏黎明 黄桂香[1] QIU Fafa;PAN Zhenzhen;LI Luanxiang;XIA Liming;HUANG Guixiang(Agricultural College,Guangxi University,Nanning 530004,Guangxi,China;Guangxi South Subtropical Agricultural Sciences Re-search Institute,Chongzuo 532400,Guangxi,China)

机构地区：[1]广西大学农学院,南宁530005 [2]广西南亚热带农业科学研究所,广西崇左532400

出　　处：《果树学报》2022年第4期631-643,共13页Journal of Fruit Science

基　　金：国家重点研发计划支持项目(2018YFD0201500);广西科技重大专项(桂科AA18118046);广西现代农业产业技术体系柑橘创新团队(nycytxgxcxtd-05-08)。

摘　　要：【目的】了解沃柑叶片响应柑橘溃疡病侵染的分子机制及易感品种与柑橘溃疡病菌的互作应答机制,筛选出柑橘溃疡病菌危害沃柑叶片时的相关应答基因,为抗性育种提供基因基础。【方法】以接种溃疡病菌后0、2、4、6和8 d的沃柑叶片为试材,利用Novaseq 6000平台进行转录组双向测序,并对数据进行生物信息学相关性分析。【结果】接种无菌水(CK)后0、2、4、6和8 d获得的Clean reads分别是48 482 127、47 270 288、50 998 549、53 201 972和47 924 731条,接种柑橘溃疡病菌(JZY)后0、2、4、6和8 d获得的Clean reads分别是51 042 967、49 552 248、46 734 029、47 940 345和45 371 891条。接种后0、2、4、6和8 d的上调差异表达基因数量分别为1、947、1081、656和2108个,下调差异表达基因数量分别为1、343、753、303和1908个。在接种后2、4、6和8 d有374个基因均表达差异,其中上调基因为61个,下调基因为313个。GO功能富集分析结果显示,沃柑叶片响应溃疡病不同时期的差异表达基因,主要集中在生物进程、分子功能和细胞组分中。KEGG注释分析结果显示,差异表达基因主要参与次生代谢物的生物合成、植物与病原体相互作用、植物激素信号转导、过氧化物酶体和蛋白质内质网合成等途径。【结论】植物与病原体相互作用、植物激素信号转导、过氧化物酶体和蛋白质内质网合成4条通路为柑橘感抗病相关的重要代谢通路。研究结果可作为深入研究柑橘种质资源感抗溃疡病基因和探究柑橘与病原菌互作分子机制的理论提供参考依据。【Objective】Orah is a hybrid with high yield, good flavor and superior quality. It is sensitive to citrus canker, caused by Xanthomonas citri subsp. citri. The mechanisms response to the susceptibility remains unknown. In this study, the transcriptome changes of Orah leaf upon the infection by X. citri subsp. citri were explored. Therefore, the molecular mechanism of citrus canker occurrence and the interaction between susceptible cultivars and X. citri subsp. citri were studied on Orah leaves. Transcriptome sequencing technology was used to screen the related response genes of citrus canker leaves, providing a genetic basis for disease resistance breeding.【Methods】Orah leaves with citrus canker were extracted from the Research Base of Agricultural College of Guangxi University to prepare citrus canker pathogen. Then the leaves with X. citri subsp. citri were inoculated by in vivo injection. The number of inoculum holes was determined by the size of the leaf until the bacterial suspension covered one side of the leaf. Centered on the veins of the leaves, one side was inoculated with sterile water as control(CK), and the other side was inoculated with citrus ulcer pathogen as experimental treatment(JZY).Samples were taken at 0, 2, 4, 6 and 8 d after inoculation. Three replicates were set for each treatment.The control sample(CK) and the treatment sample(JZY) were wrapped in tin foil, treated with liquid nitrogen and put into the refrigerator at-80 ℃ for later use. DNA was extracted by Biospin bacterial genomic DNA extraction kit, RNA was extracted by Trizol method, and transcriptome sequencing was completed by Beijing Novo Biotechnology Co., LTD. Raw data were obtained using the sequencing platform Novaseq 6000 and clean reads were obtained through quality control. The reference genome used in this experiment was the sweet orange genome. Sequencing data were analyzed and functional annotation was performed. HTSeq software was used to analyze gene expression levels. The threshold of gene expression level w

关 键 词：沃柑 柑橘溃疡病 离体接种 转录组 差异表达基因 

分 类 号：S666.1[农业科学—果树学] S436.66[农业科学—园艺学]