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棉花学报 ›› 2016, Vol. 28 ›› Issue (2): 104-114.doi: 10.11963/issn.1002-7807.201602002

• 研究与进展 • 上一篇    下一篇

3个陆地棉球蛋白基因启动子的克隆与功能初步分析

魏琦超1,2,张锐1,孙国清1,孟志刚1,周焘1,赵雷霖1,郭三堆1*   

  1. 1. 中国农业科学院生物技术研究所,北京 100081;2. 河南科技学院生命科技学院,河南 新乡 453003
  • 收稿日期:2015-05-21 出版日期:2016-03-15 发布日期:2016-03-15
  • 通讯作者: guosandui@caas.cn
  • 作者简介:魏琦超(1980―),男,博士研究生,讲师,weiqc@126.com
  • 基金资助:
    国家科技重大专项——转基因生物新品种培育(2012ZX08005-004)

Cloning and Preliminary Analysis of the Function of Three Globulin Genes Promoters from Gossypium hirsutum L.

Wei Qichao1,2, Zhang Rui1, Sun Guoqing1, Meng Zhigang1, Zhou Tao1, Zhao Leilin1, Guo Sandui1*   

  1. 1. Biotechnology Research Institute, Chinese Academy of Agricultural Sciences, Beijing 100081, China; 2. School of Life Science and Technology, Henan Institute of Science and Technology, Xinxiang, Henan 453003, China
  • Received:2015-05-21 Online:2016-03-15 Published:2016-03-15

摘要: 为克隆陆地棉来源的种子特异性启动子,根据雷蒙德氏棉测序结果,设计针对GhαGLOAGhβGLOAGhβGLOB基因编码区上游约1.5 kb序列的引物,分别以陆地棉总DNA为模板克隆了3条序列。构建了含有编码区上游序列驱动GUS的表达载体,经农杆菌介导转化野生型拟南芥。转基因拟南芥种子的GUS活性荧光检测结果表明,所克隆序列具有启动子功能,其中GhαGLOA启动子的转录活性极显著高于其他2个启动子。在转基因拟南芥成体植株的多个器官中,仅可检出痕量的GUS活性,认为所克隆启动子为种子特异性启动子。

关键词: 陆地棉; 球蛋白; 启动子; 种子特异性启动子; 顺式作用元件

Abstract: The aim of this study is to clone seed-specific promoter from Gossypium hirsutum. Using the genome sequence of Gossypium raimondii, primers were designed and synthesized. Upstream sequences of GhαGLOA, GhβGLOA, and GhβGLOB were amplified using total DNA of G. hirsutum as the template. Expression plasmids of GUS driven by the three upstream sequences were introduced into Arabidopsis thaliana. GUS activity was tested in seeds of transgenic Arabidopsis by fluorometric assay. The results showed that the three cloned sequences function as promoters, and that the promoter activity of GhαGLOA is significantly higher than that of GhβGLOA or GhβGLOB. Fluorometric analysis of GUS activity was carried out to examine the activity of these three promoters in different adult organs and tissues. Results showed that the expression level of GUS in roots, stems, leaves, and all other tissues examined, is very low, indicating that high levels of GUS activity were detected only in seeds. These results suggest that the transcriptional activity of promoters from GhαGLOA, GhβGLOA, and GhβGLOB was tightly controlled and highly seed specific.

Key words: Gossypium hirsutum L.; globulin; promoter; seed-specific promoter; cis-acting element

中图分类号: 
  • S562.03