棉花光合基因GhRCAα启动子的克隆及活性分析

摘要

In order to clone the promoter of rubisco activase gene (RCA)and understand its molecular regulation mechanism,we cloned and analyzed the cis-acting regulatory elements and activity of the 2 000 bp promoter sequence of GhRCAα from Baimian 1.The results showed that there were severalimportant cis-acting elements,including the elements responsible for light,circadian,stress,phytohormone,and other basal elements in the upstream regulatory region of GhRCAα.Further analysis of the expression pattern of GhRCAα indicated that the expression of the GhRCAα was tissue specific,which accumulated at high levels in the photosynthesis organ of leaves,but at low levels in other detected tissues,which was consistent with the results that many light-responsive and organ-specific elements were present in the promoter region of GhRCAα.Transient expression analysis in tobacco leaf showed that 2 000 bp promoter sequence of GhRCAα was able to drive the GUS expression,suggesting that the cloned promoter fragment had the activity to drive the expression of target gene.These results indicated that the GhRCAα promoter was a tissue-specific promoter,which was expected to be used for the genetic transformation of plants,and thus to better control the specific expression of important genes.%为了克隆棉花Rubisco活化酶基因(RCA)启动子,研究其表达调控的分子机制,以百棉1号为材料,对GhRCAα启动子区2 000 bp的片段进行克隆、顺式作用元件分析以及活性分析,结果表明,许多重要的顺式作用元件包括响应于光、生物钟、逆境胁迫、植物激素以及其他的基本顺式作用元件特异地存在于GhRCAα启动子区;进一步对GhRCAα进行表达特性分析发现,该基因在光合作用进行的主要位置叶片中表达量最高,在其他组织表达量很低,其表达具有组织特异性,这与该启动子区存在许多光响应及组织特异性表达相关元件的结果相一致;将克隆的GhRCAα启动子片段以烟草叶片为受体材料进行瞬时表达分析表明,GhRCAα启动子可以驱动GUS基因的表达,表明克隆的启动子片段具有驱动目标基因表达的活性.克隆的GhRCAα启动子可能是一种组织特异型启动子,有望用于植物的遗传转化,进而更好地调控重要基因的特异性表达.