【作者】 闫绍鹏；

【导师】 杨传平； 王秋玉；

【作者基本信息】 东北林业大学 ， 遗传学， 2011， 博士

【摘要】 扦插是林木无性繁殖的主要方式,主要针对一些利用种子繁殖比较困难的植物材料,以及具有优良特性的杂交种。对于一些扦插生根困难和高杂种优势的优良树种,扦插生根率低是制约利用扦插进行无性系林业发展的瓶颈。迄今对林木扦插生根的研究虽然已在生理生化的水平上取得一定进展,但扦插生根是一个复杂的生物学过程,仅从生理、生化水平进行研究并不能揭示其生根根本机理,因此需要从多个方面对扦插生根机理进行研究。本研究以欧美山杨杂种嫩枝微扦插技术研究为基础,对欧美山杨杂种扦插生根的形态解剖、生理生化进行了研究。并在此研究基础上,通过对欧美山杨杂种生根过程中不同时期的基因组进行Solexa测序、生物信息学分析,剖析插穗生根的分子机理,挖掘扦插生根过程中不定根发育相关的基因及其功能。探索生根相关基因与表型之间的相关关系,为利用基因工程育种手段改良林木提供科学依据,研究具有重要的科学意义和应用价值。本论文主要结果如下：1扦插生根过程形态解剖研究：欧美山杨杂种属于皮部诱导生根类型,根原基起源于韧皮部的薄壁细胞,愈伤组织的形成与扦插不定根的发生无直接关系,愈伤组织发达的插穗不再生根。2扦插生根过程生理学研究：IAA、ABA、ZR、GA3在欧美山杨杂种根原基的诱导和不定根的伸长阶段起着重要的作用,IAA、ZR、GA3三种激素协同参与了扦插不定根的形成过程。在扦插生根过程中根原基的诱导和形成阶段,PPO、POD含量升高,而在不定根伸长生长阶段PPO和POD含量下降。欧美山杨杂种插穗可溶性糖、蛋白质含量在扦插生根过程中总体呈下降趋势。3扦插生根的分子机理研究：插穗不定根发育是一个复杂的生物学过程,通过对欧美山杨杂种扦插生根过程中的两个时期材料进行Solexa全基因组测序,参照数字化基因表达谱差异基因检测方法筛选出表达差异基因21508个,以毛果杨基因组数据库为基础,整合多个公共数据库中杨树分子生物学资源对筛选到的差异表达基因进行相关的生物学功能注释,并列举和分析了部分可能对扦插过程中不定根发育研究有重要意义的基因。对差异表达基因进行GO分析和Pathway显著性富集分析,这些序列包括了新陈代谢、信号途径、物质运输、抗性相关、生长发育、能量代谢、细胞周期等基因。研究还获得大量功能未见报道的新基因,说明在欧美山杨杂种扦插形成不定根过程中还有许多基因水平的变化不清楚。这些研究为杨树不定根发育相关基因的克隆提供依据,也为进一步开展插穗不定根分子发育生物学研究奠定了基础。更多还原

【Abstract】 Cutting is the main form of vegetative reproduction of forest, and the key to breed quality varieties is rooting technique. For some forestry species with roots hard, low rooting cutting rate is bottleneck that restricted use of cuttings for clonal forestry development. So far the research on forests rooting have some progresses in the physiological and biochemical levels, but rooting is a complex biological process, and the study only from the physiological and biochemical level can not reveal the underlying mechanism, so we need the comprehensive study of mechanism of rooting from many aspects.Based on Populus tremula×P.tremuloides micro-cutting technology, rooting anatomical, physiological, biochemical and molecular mechanism were studied. The investigation of molecular mechanism was through on European and American roots in the different periods of Solexa genome sequencing, bioinformatics analysis, analysis of the molecular mechanism of rooting, finding related genes and their functions in the process of cutting and adventitious rooting. It also provided the scientific basis for the breeding using genetic engineering by exploring the correlation between rooting related genes and phenotypic. The research has important scientific significance. The result of this paper is as follows:1 Morphological and anatomical studies of rooting:Europe and America hybrid aspen belong to cortex inducted rooting, and root primordia was originated from the phloem parenchyma cells. The callus formation was not directly related to the occurrence of adventitious roots cutting, and the developed cutting of callus was rooted no longer.2 Research on physiology of rooting process:IAA, ABA, ZR, GA3 played an important role in the stage of induction and adventitious root primordia elongation for hybrid aspen. IAA, ZR, GA3 were involved in the collaborative formation process of adventitious roots. In the stage of root primordial induction and formation, activities of PPO, POD, IAAO were increased, while in the stage of adventitious root elongation, activities of PPO and POD were decreased. IAAO activity maintained in a high level. Soluble sugar and protein content of hybrid aspen were decreased in the process of rooting in the total.3 The molecular mechanism of cutting rooting:Adventitious root development of cuttings is a complex biological process. Through the Europe and America hybrid aspen rooting during the two periods of material conducte Solexa genome sequencing,21508 differential expression genes were selected referred to digital detection of differential gene expression profiles. Based on populus trichocarpa genome database and integrated a number of public databases in molecular biology resources, the selection of aspen to the differentially expressed genes was studied with related to biological functional annotation. Furthermore, some of the significant genes for the development of adventitious root cuttings were cited and analyzed. The genes expressed differentially were related to metabolism, signaling pathways, material transport, resistance-related, growth and development, energy metabolism, cell cycle genes and so on by conducting GO and Pathway analysis of enriched significantly. Our study also received a large number of new gene which had not been reported, indicating that the functions of many genes during the formation of adventitious roots for hybrid aspen cuttings were not clear. The research not only provided the basis for the adventitious roots of aspen clone genes, but also lay the foundation for further development of molecular biology of adventitious root cuttings.更多还原