《植物生理学报》 2012, 48(1): 67-74

甘蓝型油菜BnGOLS1基因启动子的克隆、序列分析及瞬时表达

景寅, 李煦, 汪晓峰*

北京林业大学林木育种国家工程实验室, 林木花卉遗传育种教育部重点实验室, 北京100083

通信作者：汪晓峰；E-mail: wxf801@sina.com；Tel: 010-62336114

摘　要：

以甘蓝型油菜‘德油五号’基因组DNA为模板, 通过反向PCR扩增得到肌醇半乳糖苷合成酶基因(BnGOLS1)启动子片段, 长度为827 bp。PLACE和PlantCARE启动子预测工具分析表明: 序列中含有TATA-Box、CAAT-Box等基本转录元件, 以及ABRE、DRE、HSE、w-Box等顺式作用元件。将克隆得到的BnGOLS1启动子取代pBI121中的CaMV35S启动子, 构建BnGOLS1启动子控制报告基因的GUS表达载体pBI-GS-GUS, 通过农杆菌介导的方法在油菜组织中进行瞬时表达。GUS染色结果表明BnGOLS1启动子可以驱动GUS基因在油菜组织中的表达。

关键词：油菜; BnGOLS1基因启动子; 序列分析; 瞬时表达

收稿：2011-10-14   修定：2011-11-09

资助：国家林业公益性行业科研专项(201104024)和国家林业局引进国际先进农业科学技术计划(“948”项目) (2011-4-54)。

Cloning, Sequence Analysis and Transient Expression of BnGOLS1 Gene Promoter from Brassica napus

JING Yin, LI Xu, WANG Xiao-Feng*

Key Laboratory of Genetics and Breeding in Forest Trees and Ornamental Plants of Ministry of Education, National Engineering Laboratory for Tree Breeding, Beijing Forestry University, Beijing 100083, China

Corresponding author: WANG Xiao-Feng; E-mail: wxf801@sina.com; Tel: 010-62336114

Abstract:

The BnGOLS1 promoter fragment (827 bp) was amplified from the genomic DNA of Brassica napus by inverse polymerase chain reaction. Promoter sequence analysis by PLACE and PlantCARE showed that the cloned fragment contained several putative cis-elements, such as abscisic acid response element (ABRE), dehydration-responsive element (DRE), heat shock elements (HSE), WRKY transcription factor recognition site wBox and so on, as well as TATA-Box and CAAT-Box. A recombinant vector designated as pBI-GS-GUS was generated through the replacement of CaMV35S promoter in pBI121 by the cloned BnGOLS1 promoter fragment, in which the reporter GUS is under the control of BnGOLS1 promoter. Transient expression of pBI-GSGUS in Brassica napus was performed by Agrobacterium tumefaciens mediated method. Histochemical staining of GUS revealed that the promoter of BnGOLS1 could drive the expression of GUS gene in Brassica napus.

Key words: Brassica napus; BnGOLS1 promoter; sequence analysis; transient expression