茶树花粉中丙酮酸脱氢酶（CsE1α）的定位分析及其启动子的克隆与表达

doi:10.13305/j.cnki.jts.2015.03.012

摘要/Abstract

摘要： 根据以往试验获得的CsE1α的cDNA全长序列,利用TAIL-PCR克隆CsE1α启动子。测序验证与生物信息学分析后发现,该启动子片段长336βbp,含有2个CAAT-box,2个TATA-box,2个GATA-box,1个LTR,1个G-box等顺式作用元件。构建载体转入洋葱内表皮细胞瞬时表达,启动子可启动下游报告基因,使荧光蛋白表达于整个细胞,表明所克隆的启动子具有启动功能。CsE1α与GFP融合蛋白瞬时表达表明CsE1α定位于线粒体。本实验为下一步转基因拟南芥稳定表达,进一步研究CsE1α基因的表达调控,探讨茶树花粉抗寒的分子生物学机理奠定基础。

关键词: 茶树, 丙酮酸脱氢酶, 启动子, 亚细胞定位, 抗寒

Abstract: In this paper, full-length cDNA is identified for designing the gene-special primers in TAIL-PCR to clone the promoter of CsE1α. By sequencing and bioinformatic analyzing, we observed two CAAT-box, two TATA-box, two GATA-box, one LTR and one G-box located in the 336βbp promoter region. After constructing and transferring the transient expression vectors into the onion epidermal cells, subcellular localization of CsE1α-GFP fusion proteins is verified in mitochondria, while the promoter can activate downstream gene expressing in entire cell. This experiment may provide useful information for the subsequent stable expression in transgenic Arabidopsis and the further investigation on the expression and regulation of CsE1α gene as well as the investigation on the molecular mechanism of cold resistance in pollen of Camellia sinensis.

Key words: Camellia sinensis, pyruvate dehydrogenase, promoter, subcellular localization, cold resistance

中图分类号:

杜昱林, 王伟东, 王玉花, 黎星辉. 茶树花粉中丙酮酸脱氢酶（CsE1α）的定位分析及其启动子的克隆与表达[J]. 茶叶科学, 2015, 35(3): 290-298. doi: 10.13305/j.cnki.jts.2015.03.012.

DU Yulin, WANG Weidong, WANG Yuhua, LI Xinghui. Subcellular Localization of CsE1α as well as Cloning and Expression of Its Promoter from the Pollen of Camellia sinensis[J]. Journal of Tea Science, 2015, 35(3): 290-298. doi: 10.13305/j.cnki.jts.2015.03.012.

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