植物不定根形成及其响应逆境胁迫的转录组学研究

Adventitious rooting is an ecologically and economically important developmental process. The formation of adventitious roots is one of the most important mechanisms of vegetative propagation in plants and one of the most important methods for the commercial production of horticultural species. In ecosystems in which soil disturbance is frequent, adventitious rooting can be an alternative or supplement to seed propagation; it is an important plant response to environmental stresses and is a strategy for plant propagation under stress. The formation of adventitious roots has been associated with an important aspect of tissue dedifferentiation involving the shifting of cells from normal morphogenetic pathways to functions associated with the development of root primordia. Recently, many researchers have revealed large number of genes that specifically regulate adventitious rooting at molecular level in plants. The microarray analyses have been used to investigate the gene expression changes involved in adventitious root formation; however, the transcriptomic study on the global gene expression profiles during adventitious rooting is still limited and require further investigated..In this research, the transcriptome approach based on RNA-Seq technology will be used to exhibit gene expression profiles during adventitious rooting in mung bean [Vigna radiata (L.) Wilczek] and in response of adventitious rooting to salt, osmotic, and heavy metal stresses. The objective of this study is to reveal the global genomic mechanisms underlying adventitious rooting and its response to abiotic stresses. Our aims are: (1) to gain the transcriptomic data of mung bean, (2) to investigate the changes of global gene expressions, the regulation of the gene expression, and the regulation mechanisms, during the induction stage and initiation stage of adventitious root; (3) to explore the roles of auxin IBA and signaling molecules NO and H2O2 on global gene expression, and the molecule mechanism regulating adventitious rooting; (4) to reveal the gene expression changes during adventitious rooting under salt, osmotic, and heavy metal stresses, and the molecular mechanisms in response to the stresses. The answers of these issues will be scientific important for illuminating the basic theory problems such as plant cell dedifferentiation and organogenesis, and the molecular mechanisms of plant in response to stresses. Also, it will be practical importance for further studying on mung bean genome, functional genes, genetic breeding, as well as seedling propagation of difficulty-to root species.

不定根是植物无性繁殖的重要途径和园林苗木生产的重要方式，是在环境胁迫下植物的繁殖策略和对种子繁殖的补充，具有重要的经济和生态意义。不定根的形成也是研究植物细胞脱分化和再分化的实验模型。在全基因层面开展不定根形成及其响应胁迫的分子机制的研究还很少。该项目运用基于RNA-Seq技术的转录学组方法，以绿豆为材料，研究植物不定根形成及其响应盐、渗透和重金属胁迫的基因表达变化，在全基因层面探讨植物不定根形成及其响应逆境胁迫的分子机制，揭示: (1)绿豆转录组，(2)植物不定根诱导和起始阶段基因表达变化、调控及其机制，(3)IBA和信号分子H2O2和NO对基因表达的影响及其调控不定根形成的分子机制， (4)逆境胁迫下不定根形成过程中基因表达的变化及其响应胁迫的分子机制。探讨植物细胞脱分化和响应逆境胁迫的分子机制等重要科学问题，促进绿豆基因组、基因资源、遗传育种和难生根植物种苗繁育等领域研究的开展。

以绿豆为材料，建立了绿豆幼苗下胚轴不定根形成实验模型，运用转录组和qRT-PCR技术对不定根诱导和起始两个阶段的基因表达，外源生长素和信号分子H2O2和NO促进不定根形成的基因表达进行了研究，探讨了干旱(渗透)和重金属胁迫下绿豆幼苗不定根形成及其响应机制。通过De Novo方法组装了78697个绿豆Unigene，注释29029个基因，建立了绿豆转录组数据库，在不定根诱导阶段和起始阶段分别鉴定到11717和12737个差异表达基因。IBA显著促进不定根形成，提高不定根诱导和起始阶段表达基因的数量，在不定根诱导阶段上调基因表达，在不定根起始阶段下调基因表达。IBA显著影响生长素信号途径、核糖体装配与蛋白质合成、光合作用、氧化还原活性和次生细胞壁合成等功能，显著影响核糖体生物发生、植物激素信号转导、戊糖和已糖转化、光合作用、苯丙氨酸合成、萜类物质合成、黄酮化合物合成和苯基丙氨酸合成等代谢通路。H2O2促进不定根形成，增加了不定根形成两个阶段表达基因的数量，上调基因表达。显著影响参与胁迫应答、细胞氧还平衡、氧化胁迫应答、细胞壁松弛与修饰、代谢过程、转录调控因子、植物激素信号通路等基因，特别是编码HSPs和HSFs的基因被显著上调。NO促进不定根形成，NO供休SNP增加了不定根诱导阶段表达基因的数量，降低了不定根起始阶段表达基因的数量。NO显著调控与氮化合物应答、胁迫应答、氧化胁迫应答、细胞壁修饰、信号转导、蛋白加工、次生代谢、转录因子，以及植物激素信号通路等相关基因的表达。Cd和山梨醇胁迫显著影响细胞内氧化还系统平衡，增加膜脂过氧化，抑制不定根的形成。IBA和NO都能够减轻两种胁迫对植物不定根的抑制，其作用机制是调控细胞内氧化还原系统状态，减缓胁迫造成的膜脂过氧化，提高细胞抗逆能力。NO和生长素通过相同的途径调控植物细胞对胁迫的响应。该项研究初步阐明了植物不定根形成及其响应胁迫的机制，为植物细胞脱分化与再分化，生长素和信号分子H2O2和NO等的生物学机制，以及绿豆生物学和遗传学研究，积累了重要科学资料。