OsAOS2基因在水稻抗稻瘟病菌和白叶枯病菌中的作用  

Effects of OsAOS2 gene on rice resistances to Magnaporthe oryzae and Xanthomonas oryzae pv.oryzae

作　　者：周永林 庄小倩 王朝露 林巧霞 李正康 许丹洁 陈晓婷 ZHOU Yonglin;ZHUANG Xiaoqian;WANG Zhaolu;LIN Qiaoxia;LIZhengkang;XU Danjie;CHEN Xiaoting(College of Life Sciences,Fujian Agriculture and Forestry University/Key Laboratory of Biopesticide and Chemical Biology,Ministry of Education/Fujian Educational Department Key Laboratory for Plant and Microbe Interaction,Fuzhou,Fujian 350002,China)

机构地区：[1]福建农林大学生命科学学院/生物农药与化学生物学教育部重点实验室/福建省教育厅植物微生物互作重点实验室,福建福州350002

出　　处：《福建农林大学学报(自然科学版)》2025年第2期145-156,共12页Journal of Fujian Agriculture and Forestry University:Natural Science Edition

基　　金：福建省自然科学基金面上项目(2022J01140);福建省科技重大专项(2022NZ03004);福建农林大学科技创新专项(KFA17308A、CXZX2020152D)。

摘　　要：【目的】研究OsAOS2在水稻抵抗稻瘟病菌和白叶枯病菌中的作用,为水稻抗病育种工作提供依据。【方法】采用同源重组方法和CRISPR/Cas9系统分别构建OsAOS2的过表达载体和敲除载体,遗传转化获得转基因水稻,筛选得到过表达株系(OsAOS2-OE)和纯合敲除株系(OsAOS2-KO)。将野生型水稻日本晴(NPB)植株分别接种稻瘟病菌和白叶枯病菌后,通过qRT-PCR检测OsAOS2表达量的变化情况;将OsAOS2转基因水稻和NPB分别接种稻瘟病菌和白叶枯病菌后,观察植株表型的变化;在稻瘟病菌侵染下,对OsAOS2转基因水稻和NPB进行抗病相关基因的qRT-PCR分析;在几丁质和flg22诱导下,检测水稻活性氧(ROS)积累情况;同时,分析接种稻瘟病菌24 h后OsAOS2-KO和NPB的转录组数据。【结果】qRT-PCR分析表明,NPB接种稻瘟病菌48 h和白叶枯病菌8 d时,OsAOS2表达量显著上调。对OsAOS2转基因水稻和NPB喷雾接种稻瘟病菌7 d后,NPB病斑面积显著大于OsAOS2-OE且小于OsAOS2-KO;剪叶法接种白叶枯病菌14 d后,NPB病斑面积显著小于OsAOS2-KO。对OsAOS2转基因水稻接种稻瘟病菌后,抗病相关基因OsPBZ1、OsPR1a、OsPAL1、OsLOX5在OsAOS2-OE中的表达量显著上调。在几丁质和flg22诱导下,OsAOS2-KO中ROS的积累量显著低于NPB,但是起峰时间比NPB有所提前。转录组数据分析表明,上调基因和下调基因分别有1 605个和1 151个。上调差异基因主要富集在金属离子结合和核糖体通路,下调差异基因主要富集在翻译通路和细胞外区域。韦恩图显示:52个基因在NPB中受稻瘟病菌诱导,但在OsAOS2-KO中受到抑制;11个基因在NPB中受抑制,但在OsAOS2-KO中受诱导。对这63个表达模式相反的差异基因进行GO和KEGG分析发现,参与应激反应和植物激素信号转导通路的基因数最多。【结论】稻瘟病菌和白叶枯病菌可以诱导OsAOS2的表达,OsAOS2通过病原菌分子模式触发的免疫(PTI)途径和茉莉酸(JA)、水杨酸(SA)【Objective】The effects of OsAOS2 on rice resistances to Magnaporthe oryzae and Xanthomonas oryzae pv.oryzae(Xoo)were studied in order to provide basis for rice disease resistance breeding.【Method】OsAOS2 overexpression vector and knockout vector were constructed by homologous recombination method and CRISPR-Cas9 system,respectively.Transgenic rice lines were obtained by genetic transformation,and overexpression lines(OsAOS2-OE)and homozygous knockout lines(OsAOS2-KO)were identified.The expression profiles of OsAOS2 was detected by qRT-PCR after inoculation with M.oryzae and Xoo.The phenotypes of OsAOS2 transgenic rice and wild type Nipponbare(NPB)after inoculation with M.oryzae and Xoowere detected.The qRT-PCR analyses of defense-related genes were performed under the infection of M.oryzae.The accumulation of reactive oxygen species(ROS)inOsAOS2 transgenic rice was detected under the induction of chitin and flg22.Meanwhile,the transcriptome data of OsAOS2-KO and NPB 24 h after inoculation with M.oryzae was analysed.【Result】qRT-PCR analyses showed that OsAOS2 expression was significantly up-regulated at 48 h and 8 d after inoculation with M.oryzae and Xoo,respectively.7 days post inoculation of OsAOS2 transgenic rice and NPB with M.oryzae showed that the lesion area of NPB was significantly larger than that of OsAOS2-OE and smaller than that of OsAOS2-KO;14days postinoculation of the above rice materials with Xoo showed that the lesion length of NPB was significantly smaller than that of OsAOS2-KO;and the expression patterns of defense related-genes were examined in OsAOS2 transgenic rice inoculated with M.oryzae,and it was found that the defense related-genes,OsPBZ1,OsPR1a,OsPAL1,and OsLOX5,were significantly upregulated in OsAOS2-OE transgenic rice.The ROS accumulation in OsAOS2-KO was significantly lower than that in the wild type induced by chitin and flg22,and the peak time of ROS in OsAOS2-KO was slightly earlier than NPB.The transcriptome data of OsAOS2-knockout transgenic rice inoculated with M.

关 键 词：OsAOS2 茉莉酸 抗病性 稻瘟病 白叶枯病 转录组分析 

分 类 号：S511[农业科学—作物学]