《植物生理学报》 2016, 52(10): 1505-1511

拟南芥花粉特异性复合式启动子的构建及活性分析

刘石娟^*, 王雪

曲阜师范大学生命科学学院, 曲阜273165

通信作者：刘石娟；E-mail: sjliu@mail.qfnu.edu.cn

摘　要：

组织特异性启动子在植物基因工程中具有重要作用, 利用组织特异性调控元件或增强子序列构建复合式启动子是获得高活性组织特异性启动子的重要途径。在本研究中, 将拟南芥泛素C端水解酶基因(AtUCH4)的增强子序列与其同源启动子融合, 构建成复合式启动子并与GUS报告基因连接。GUS组织化学染色和酶活分析表明该启动子能够在转基因拟南芥花粉中特异性高效表达, 其活性远高于AtUCH4启动子。定量逆转录PCR (qRT-PCR)实验结果表明融合启动子的活性是AtUCH4启动子的3.3倍, 进一步证实该融合启动子是一个花粉特异性高表达启动子, 从而可为作物遗传改良提供新的候选组织特异性启动子。

关键词：启动子; 顺式调控元件; 组织特异性; 花粉

收稿：2016-07-22   修定：2016-09-05

资助：国家自然科学基金(31670270和31200196)和山东省自然科学基金(ZR2011CQ016)。

Construction and functional analysis of a synthetic pollen-specific promoter in Arabidopsis thaliana

LIU Shi-Juan^*, WANG Xue

College of Life Sciences, Qufu Normal University, Qufu, Shandong 273165, China

Corresponding author: LIU Shi-Juan; E-mail: sjliu@mail.qfnu.edu.cn

Abstract:

Tissue-specific promoters have important roles in plant genetic engineering. Highly expressing tissue-specific promoters can be obtained by synergistic combinations of multiple transcriptional activator elements. In this study, the enhancer fragments of Arabidopsis thaliana ubiquitin C-terminal hydrolase gene AtUCH4 have been amplified by PCR, cloned and fused to the homologous promoter sequence together with GUS. GUS histochemical staining and fluorometric analysis show that this synthetic promoter was highly expressed in pollen of the transgenic A. thaliana plants, and its activity was much higher than that of the AtUCH4 promoter. QRT-PCR experiments further confirmed that this fusion promoter was a highly pollen-specific promoter, which provides a new candidate tissue-specific promoter for genetic improvement of crops.

Key words: promoter; cis regulatory element; tissue-specific; pollen