杨树中MAX2同源基因的克隆

发布时间：2018-04-15 15:19

本文选题：植物分枝 + 独脚金内酯　；参考：《华中农业大学》2016年硕士论文

【摘要】：分枝为植物非常复杂的生理发育过程,涉及到多种生理机制,植物激素对分枝的影响是其中重要的一环。2008年发现了一类新型植物激素——独脚金内酯(strigolactone, SLs),其功能为参与调控植物地上部分分枝。当前,关于独脚金内酯调控分枝的机理机制研究已取得一定进展,在拟南芥(Arabi-dopsis thaliana)、水稻(Oryza sativa)、豌豆(Pisum sativum L.)、矮牵牛(petunia hybrida)等物种中已克隆出许多与独脚金内酯相关的基因。但至今独脚金内酯的合成和信号传导的具体途径尚不清晰,在不同物种中已克隆的基因亦有限,特别是其在木本植物中的作用机制还处于起步阶段。而杨树作为木本植物中的模式植物,探究独脚金内酯对其分枝机理的具影响有重要意义。本实验通过生物信息学方法,调取11个拟南芥与独脚金内酯合成及信号转导相关的基因,比对出其在毛果杨(Populus trichocarpa)中对应的同源基因,对筛选出的毛果杨独脚金内酯各途径的同源基因,分析各基因的功能及表达情况,挑选出拟南芥及毛果杨独脚金内酯信号转导途径中MAX2的同源基因为目的基因进行克隆。目前,实验部分使用Gateway方法已克隆出AtMAX2基因(AT2G42620),同时将AtMAX2基因在Col-0型拟南芥中过量表达；同时,以欧洲山杨×银白杨(Populus tre mula×P. Alba INRA clone N717 1-B4)组培苗为植物材料,将AtMAX2在717杨树中过量表达；克隆出杨树中MAX2的同源基因PtrMAX2c (Potri.006G068500).构建已转化AtMAX2到717杨,已鉴定出阳性植株；构建已转化PtrMAX2c基因到Col-0型拟南芥中,已得到PtrMAX2c转AT T1代种子；经过3代拟南芥植株的阳性苗筛选及RT-PCR表达量鉴定,已得到AtMAX2转Col-0型拟南芥过量表达转基因植株的纯合子株系9个；通过阳性苗筛选筛选及RT-PCR表达量鉴定,筛选出At-MAX2转717杨树过量表达株系3个。
[Abstract]:Branching is a very complex process of plant physiological development, which involves a variety of physiological mechanisms.The effect of phytohormone on branching is an important one. In 2008, a new kind of plant hormone, strigolactone, SLS, was found, which is involved in the regulation of shoot branching of plants.At present, some progress has been made on the mechanism of the regulation of branching by monogolide. In Arabidopsis thaliana, Oryza sativaa, Pisum sativum L., petunia hybrida), many genes related to monopodide have been cloned in Arabidopsis thaliana.However, the specific pathway of the synthesis and signal transduction of goldactone is not clear, and the cloned genes in different species are limited, especially its mechanism of action in woody plants is still in its infancy.Poplar, as a model plant in woody plants, is of great significance to explore the effect of diptyrolactone on its branching mechanism.In this study, 11 genes related to the synthesis and signal transduction of goldactone in Arabidopsis thaliana were obtained by bioinformatics, and their homologous genes in Populus trichocarpa were compared.The homologous genes of MAX2 in the signal transduction pathway of Poplar monopodide from Arabidopsis thaliana and Populus tomentosa were selected for cloning by analyzing the function and expression of these genes.At present, the AtMAX2 gene AT2G42620 was cloned by Gateway method, and the AtMAX2 gene was overexpressed in Col-0 type Arabidopsis thaliana. At the same time, the plantlets of Populus tre mula 脳 P. INRA clone N7171-B4 were used as plant materials.AtMAX2 was overexpressed in 717 poplar and the homologous gene PtrMAX2c Potri.006G068500 was cloned.Positive plants were identified by construction of transformed AtMAX2 to 717 poplar, transformed PtrMAX2c gene into Col-0 type Arabidopsis thaliana, and transgenic AT T1 seeds were obtained. After 3 generations of Arabidopsis thaliana plants, the positive seedlings were screened and RT-PCR expression was identified.Nine homozygous lines of Col-0 type Arabidopsis thaliana transgenic plants with AtMAX2 were obtained, and 3 lines of At-MAX2 transferred 717 poplar were screened by screening positive seedlings and identifying RT-PCR expression.
【学位授予单位】：华中农业大学
【学位级别】：硕士
【学位授予年份】：2016
【分类号】：S792.11

【参考文献】