中棉光诱导基因Gacab启动子的克隆及其功能分析  

Cloning and Functional Analysis of the Light-inducible Gossypium arboreum Gacab Promoter

作　　者：李为民[1] 王志兴[1] 裴新梧[1] 贾士荣[1] 

机构地区：[1]中国农业科学院生物技术研究所,北京100081

出　　处：《农业生物技术学报》2004年第3期253-257,共5页Journal of Agricultural Biotechnology

摘　　要：利用加接头法,从金华中棉(Gossypiumarboreum)中分离获得了捕光叶绿素a/b蛋白复合体基因Gacab编码区5'上游的调控序列1009bp,命名为GacabP,与公布的其它植物cab启动子序列比较,没有明显的同源性。将GacabP和197、504、779bp的5'端缺失体分别与gus(uidA)基因融合,构建植物表达载体。用GacabP::gus转化烟草(Nicotianatabacumcv.NC89),GUS组织化学分析发现GacabP驱动gus基因在转基因烟草的叶片表达。在暗培养的转基因烟草叶片中GacabP不表现活性,而光能够诱导gus基因的表达,表明GacabP是一种光诱导型启动子。用基因枪轰击水稻愈伤组织,结果显示,GacabP及3种不同长度的缺失体均可驱动gus基因的瞬时表达,以504bp的活性最高,瞬时表达水平明显高于35S启动子,197、779和1009bp的表达减弱,由此推测-197^-1bp为Gacab基因的基本启动子,-504^-197bp间包含正调控元件,-1009^-504bp间包含负调控元件。A 1 009 bp promoter sequence of cab gene that encodes chlorophyll a/b binding protein belonging to a class of light-inducible proteins was cloned from Gossypium arboreum . Sequence analysis showed no obvious homology to previous published cab promoters. The full length Gacab P and 5' end deletions with a length of 197, 504 and 779 bp were fused with gus (uid A) gene and plant expression vectors were constructed respectively. The construct with Gacab P :: gus was used for transformation of Nicotiana tabacum cv.NC89 and transgenic tobacco plants generated. GUS histochemical assay showed that GUS was specifically expressed in leaves and young green tissues. GUS was not detected in leaves of transgenic plants grown in the dark for 6 d, whereas GUS was highly expressed in leaves of these plants further induced with light for 6 d, demonstrating Gacab P was a light-inducible promoter. The transient GUS expression in rice calli indicated the expression level of 504 bp was the highest, which was stronger than that of the 35S promoter, while expression was reduced for 197, 779 and 1 009 bp. It suggests that -197 ~ -1 bp is a basic promoter of Gacab, some positive regulatory elements exist in -504 ~ -197 bp, and fragment -1 009 ~ -504 bp may contain negative elements.

关 键 词：植物 金华中棉 光诱导基因 Gacab启动子 克隆 调控元件 

分 类 号：Q943.2[生物学—植物学]