柑橘转录因子CsMYB77与CsMYB21调控果实成熟的功能分析

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目前,我国柑橘产业亟待解决的问题之一是果实熟期集中,早、晚熟品种少,熟期结构的不合理严重降低了我国柑橘的市场竞争力。如何通过分子手段调控柑橘果实熟期,解析果实成熟机理对发展柑橘产业意义重大。前人通过对奉节脐橙及其晚熟芽变进行转录组测序,筛选到与果实成熟相关的转录因子CsMYB77和CsMYB21,将这两个转录因子转入番茄中进行功能验证。本研究在前人研究的基础上,对T1和T2代超表达CsMYB77和CsMYB21的番茄果实进行功能验证,确定了转录因子在果实成熟过程中的功能。主要研究结果如下:  1、对T1代超表系番茄进行表型观察与统计,发现与野生型(WT)和空载对照(Control)相比,超表系果实的破色时间晚3-6天。对所有超表达CsMYB77和CsMYB21的株系进行目的基因表达分析,各株系选择一株果实种子进行T2代播种。  2、对T2代超表系番茄进行表型观察与统计,与Control相比,超表系果实的破色天数都显著变晚,果实整体的破色时间向后延迟3-6天,与WT相比,只有MYB771-3一个超表系果实的破色天数显著晚于野生型。对T2代番茄果实进行成熟相关的生理指标检测,乙烯释放速率检测结果显示,野生型和空载对照果实在34DAF时乙烯释放速率开始增加,在39DAF达到释放高峰,而超表系在39DAF时乙烯释放速率才开始增加,在42DAF或者45DAF达到释放高峰。高效液相色谱检测果实中番茄红素的结果显示,26DAF时野生型、空载对照和超表系果实中均没有检测到番茄红素的存在,36DAF时野生型和对照果实中番茄红素开始积累,超表系在39DAF时才开始有番茄红素的积累。以上结果说明,超表达CsMYB77和CsMYB21的番茄果实较野生型和空载对照果实破色变晚,即延迟了果实的成熟进程。  3、对成熟相关基因进行表达分析发现,乙烯合成基因SlACS4和SlACO1,八氢番茄红素合成基因SlPSY1,成熟调控转录因子SlRIN和SlNOR的表达水平在野生型和空载对照中从36DAF开始增加。超表系中,39DAF时这些成熟相关基因的表达才开始增长,这与之前果实着色的统计结果和生理指标的测定结果一致,说明超表达CsMYB77和CsMYB21的番茄果实能够延迟成熟3天左右。
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