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棉花学报  2011 

棉花种子特异性启动子的克隆及表达载体构建

DOI: 1002-7807(2011)05-0427-06, PP. 427-432

Keywords: Lea蛋白,D113基因启动子,D34基因启动子,启动子分析,表达载体

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Abstract:

种子特异性表达启动子是植物种子基因工程改良的重要工具。Lea(Lateembryogenesisabundant)蛋白是胚胎发育后期种子中大量积累的一系列蛋白质,因此,其调控序列可能提供一个很好来源的种子特异性表达启动子。为研究植物Lea蛋白基因启动子在种子中的特异性表达,本研究通过PCR扩增,从亚洲棉(GossypiumarboreumL.)石系亚1号中克隆了Lea蛋白基因家族中D113基因上游1262bp的调控序列和D34基因上游1445bp的调控序列。DNA序列分析结果表明,克隆到的两个片段与已报道的Lea蛋白基因同一家族该基因的对应序列的相似性分别达到97.43%和94.27%。分别用限制性内切酶HindⅢ和xbaI双酶切重组质粒pGEMT-D和改造后的表达载体pBI121-T,分别回收pGEMT-D重组质粒中的小片段和pBI121-T植物表达载体中的大片段,经连接、转化和鉴定,获得由D113基因启动子和D34基因启动子驱动报告基因GUS的新型植物表达载体pBI121-D113、pBI121-D34,为外源基因在棉花种子中的定位表达研究奠定基础。

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