【作者】 刘斌；

【导师】 任华中；

【作者基本信息】 中国农业大学 ， 蔬菜学， 2017， 博士

【摘要】 植物激素赤霉素(GA)是高等植物体内的一类四环二萜羧酸化合物,在植物各个生长发育阶段起着至关重要的作用。前人已在赤霉素促进种子发芽,调控植物营养生长和生殖生长等方面开展了较多研究,但有关其调控植物果实发育方面的分子机制尚不明确。本文以黄瓜为试材,通过时空表达、功能验证和表达谱分析,研究了 GA受体蛋白CsGID1a对黄瓜果实心室形成的调控机制,旨在丰富黄瓜果实发育的理论基础,取得的主要结果如下:(1)黄瓜CsGID1a和CsGIID1b属于高等植物赤霉素受体基因GID1家族的同源基因,均含有受体基因所特有的HGG和GXSXG两个保守域,在黄瓜的茎尖和根尖处高度表达,外源GA3处理可导致反馈调节,使CsGID1a和CsGID1b的表达量下调,表明CsGID1a和CsGID1b均具备赤霉素受体基因的典型特征。(2)进化树分析表明CsGID1a处于双子叶植物GID1主效基因ac族,时空表达分析显示CsGID1a在根尖、茎尖、雌花芽及果实中的表达显著高于CsGID1b,干扰CsGID1b表达并不会影响植株生长,表明CsGID1a可能是主效基因。(3)通过原位杂交和GUS染色在花和果实中对CsGID1a的表达进行更详细分析表明,CsGID1a在黄瓜雄蕊、雌蕊及果实的心室壁上特异表达。通过黄瓜遗传转化获得的CsGID1a干扰转基因植株,与对照相比,干扰植株明显矮化,果实心室发育不完全。表达谱分析表明在干扰转基因植株中,生长素合成与运输基因和细胞周期相关基因的表达量都显著低于对照组;同时激素测定与细胞切片观察的结果表明,CsGID1a干扰转基因植株果实中生长素的分布和果实细胞分化受到影响。(4)在拟南芥中的功能验证发现,异源表达CsGID1a可部分恢复双突变体gid1agid1c的植株高度以及植物育性表型,同时也对果实心室的发育造成了影响,部分拟南芥转基因株系的果实还形成了类似黄瓜果实的三心室。更多还原

【Abstract】 Gibberellins(GAs)are phytohormones with many roles in plant growth,including the regulation of seed germination,vegetative growth and reproductive development.However,little is known about the relationship between GA perception and fleshy fruit ontogeny,and particularly locule formation.In this study,cucumber(Cucumis sativus L.),a typical monoecious plant that produces fleshy fruit,was used as an experimental system to address this question.The main results are as follows:(1)CsGID1a and CsGID1b are two homologues of Arabidopsis.thaliana GID1 genes.An amino acid alignment revealed that CsGID1a and CsGID1b proteins have the conserved HSL motifs HGG and GXSXG.GA3 treatment caused a feedback regulation of CsGID1a and CsGIDlb.(2)Phylogenetic analysis revealed that CsGID1a and CsGID1b aligned with the GID1ac and GID1b groups,respectively.CsGID1a transcripts were expressed at far higher levels than those of CsGID1b in female flower buds and fruits,so we hypothesized that,of the two genes,CsGID1a is the more likely to play the predominant role in fruit development.This idea was supported by an RNAi experiment revealing that RNAi lines with significantly reduced CsGID1b transcript levels,in which the expression of CsGIDla was not altered compared with the control,showed normal height and fruit development.(3)The expression pattern of CsGID1a was found to be closely correlated with fruit locule formation by in situ hybridization and a promoter::GUS(β-glucuronidase)assay,and silencing CsGID1a in cucumber resulted in fruits with abnormal carpels and locules.The DGE analysis suggested that expression of genes related to auxin synthesis and transport,as well as the cell cycle,was altered in CsGID1a-RNAi fruits;a result that was supported by comparing the auxin content and cellular structures of the control and transgenic fruits.(4)Over-expression of CsGID1a in the Arabidopsis thaliana double mutant(gid1a gid1c)partial rescued of the semi-dwarfed phenotype and resulted in ’cucumber locule-like’ fruits.更多还原