【目的】探究文心兰miR168的分子进化特性,揭示文心兰miR168a及其候选靶基因响应软腐病菌侵染的表达模式。【方法】以文心兰miRNA文库、转录组数据库及miRbase数据库获取的植物miR168前体和成熟体序列为研究对象,利用生物信息学方法对...【目的】探究文心兰miR168的分子进化特性,揭示文心兰miR168a及其候选靶基因响应软腐病菌侵染的表达模式。【方法】以文心兰miRNA文库、转录组数据库及miRbase数据库获取的植物miR168前体和成熟体序列为研究对象,利用生物信息学方法对其进行系统进化树构建、保守性分析、前体二级结构分析及靶基因预测;通过PCR技术克隆文心兰miR168a前体,采用qRT-PCR技术分析miR168a及其候选靶基因在文心兰不同组织(根、叶、假鳞茎)和感染软腐病(0,12,24,36和48 h)不同时间的表达模式。【结果】植物miR168序列保守性较高,来源于miR168前体的5p臂保守性高于3p臂,miR168前体分歧较大,茎序列的保守性高于环序列;从文心兰中克隆得到93 bp miR168a前体序列,包含21 nt miR168a成熟体序列。进化分析表明,物种亲缘关系对其前体和成熟体序列的形成影响较小;二级结构显示,文心兰miR168a前体能形成典型的发卡结构,其成熟体位于前体的5p臂;随着软腐病侵染时间延长,文心兰miR168a与其候选靶标RNA剪切复合体核心蛋白(AGO1)、RPM1互作蛋白(RIN4)、羧酸酯酶(CXE8)的表达趋势基本呈负相关,而与其候选靶标RNA解旋酶(PRH47A)、泛素羧基末端水解酶(UCH)的表达趋势呈正相关,说明可能存在其他miR168家族成员或miRNAs同时调控。【结论】文心兰miR168与其他植物miR168进化上兼具保守性和特异性,miR168a上调表达可能会增强文心兰的抗病性。展开更多
[Objective]The paper was to study the physiological mechanism of Oncidium hybridum‘Gower Ramsey’infected by Erwinia carotovora pv.carotovora.[Method]O.hybridum‘Gower Ramsey’was inoculated with E.carotovora by need...[Objective]The paper was to study the physiological mechanism of Oncidium hybridum‘Gower Ramsey’infected by Erwinia carotovora pv.carotovora.[Method]O.hybridum‘Gower Ramsey’was inoculated with E.carotovora by needle-punching method.The leaves inoculated with sterile water were used as control,and samples were collected at 12,24,36 and 48 h post inoculations to determine the physiological indexes,respectively.[Result]The contents of MDA,flavonoid,lignin,soluble protein and the activities of POD,SOD,CAT,PPO and PAL in O.hybridum leaves first increased then decreased.At 12 h post inoculation,the activities of SOD and CAT increased significantly by 40.49%and 37.77%(P<0.01,the same below),respectively;the contents of flavonoid increased significantly by 21.56%(P<0.05,the same below);and the activities of PAL,POD and PPO increased significantly by 42.29%,81.8%and 37.98%,respectively.The contents of MDA and lignin increased significantly by 57.98%and 58.72%at 24 h,respectively;the content of soluble protein increased by 87.71%at 24 h,and 61.28%at 36 h,while the content of Pro increased at 24 h(P>0.05,the same below),and increased significantly by 23.66%at 36 h;the content of soluble sugar content had no significant difference compared with the control.Additionally,the chlorophyll a and carotenoid contents tended to decrease dramatically,while chlorophyll b increased significantly by 95.33%at 24 h,then decreased.The initial fluorescence parameter of Fm/Fv and Y(II)decreased significantly by 47.14%and 54.61%,respectively,while the Y(NO)increased significantly by 33.61%,compared with the control.[Conclusion]The contents of Pro,MDA,lignin,soluble protein and the activities of POD,SOD,CAT,PPO and PAL in O.hybridum leaves are closely related to the infection of E.carotovora,which provide new clues for revealing the infection mechanism of E.carotovora in O.hybridum.展开更多
文摘【目的】探究文心兰miR168的分子进化特性,揭示文心兰miR168a及其候选靶基因响应软腐病菌侵染的表达模式。【方法】以文心兰miRNA文库、转录组数据库及miRbase数据库获取的植物miR168前体和成熟体序列为研究对象,利用生物信息学方法对其进行系统进化树构建、保守性分析、前体二级结构分析及靶基因预测;通过PCR技术克隆文心兰miR168a前体,采用qRT-PCR技术分析miR168a及其候选靶基因在文心兰不同组织(根、叶、假鳞茎)和感染软腐病(0,12,24,36和48 h)不同时间的表达模式。【结果】植物miR168序列保守性较高,来源于miR168前体的5p臂保守性高于3p臂,miR168前体分歧较大,茎序列的保守性高于环序列;从文心兰中克隆得到93 bp miR168a前体序列,包含21 nt miR168a成熟体序列。进化分析表明,物种亲缘关系对其前体和成熟体序列的形成影响较小;二级结构显示,文心兰miR168a前体能形成典型的发卡结构,其成熟体位于前体的5p臂;随着软腐病侵染时间延长,文心兰miR168a与其候选靶标RNA剪切复合体核心蛋白(AGO1)、RPM1互作蛋白(RIN4)、羧酸酯酶(CXE8)的表达趋势基本呈负相关,而与其候选靶标RNA解旋酶(PRH47A)、泛素羧基末端水解酶(UCH)的表达趋势呈正相关,说明可能存在其他miR168家族成员或miRNAs同时调控。【结论】文心兰miR168与其他植物miR168进化上兼具保守性和特异性,miR168a上调表达可能会增强文心兰的抗病性。
基金Young and Middle-aged Teacher Educational Research Project of Fujian Province(KLA19009A)&Major Agricultural Science and Technology Project of Fujian Province(2015NZ0002-1)&Science and Technology Innovation Fund of Fujian Agriculture and Forestry University(KF2015108).
文摘[Objective]The paper was to study the physiological mechanism of Oncidium hybridum‘Gower Ramsey’infected by Erwinia carotovora pv.carotovora.[Method]O.hybridum‘Gower Ramsey’was inoculated with E.carotovora by needle-punching method.The leaves inoculated with sterile water were used as control,and samples were collected at 12,24,36 and 48 h post inoculations to determine the physiological indexes,respectively.[Result]The contents of MDA,flavonoid,lignin,soluble protein and the activities of POD,SOD,CAT,PPO and PAL in O.hybridum leaves first increased then decreased.At 12 h post inoculation,the activities of SOD and CAT increased significantly by 40.49%and 37.77%(P<0.01,the same below),respectively;the contents of flavonoid increased significantly by 21.56%(P<0.05,the same below);and the activities of PAL,POD and PPO increased significantly by 42.29%,81.8%and 37.98%,respectively.The contents of MDA and lignin increased significantly by 57.98%and 58.72%at 24 h,respectively;the content of soluble protein increased by 87.71%at 24 h,and 61.28%at 36 h,while the content of Pro increased at 24 h(P>0.05,the same below),and increased significantly by 23.66%at 36 h;the content of soluble sugar content had no significant difference compared with the control.Additionally,the chlorophyll a and carotenoid contents tended to decrease dramatically,while chlorophyll b increased significantly by 95.33%at 24 h,then decreased.The initial fluorescence parameter of Fm/Fv and Y(II)decreased significantly by 47.14%and 54.61%,respectively,while the Y(NO)increased significantly by 33.61%,compared with the control.[Conclusion]The contents of Pro,MDA,lignin,soluble protein and the activities of POD,SOD,CAT,PPO and PAL in O.hybridum leaves are closely related to the infection of E.carotovora,which provide new clues for revealing the infection mechanism of E.carotovora in O.hybridum.
