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作物学报 ›› 2017, Vol. 43 ›› Issue (08): 1161-1169.doi: 10.3724/SP.J.1006.2017.01161

• 作物遗传育种·种质资源·分子遗传学 • 上一篇    下一篇

大豆转录因子基因GmNF-YCa可提高转基因拟南芥渗透胁迫的耐性

李敏1,2,于太飞1,2,徐兆师2,张双喜3,闵东红1,陈明2,马有志2,柴守诚1,*,郑炜君1,*   

  1. 1西北农林科技大学农学院 / 旱区作物逆境生物学国家重点实验室, 陕西杨凌 712100; 2中国农业科学院作物科学研究所 / 国家农作物基因资源与基因改良重大科学工程 / 农业部麦类生物学与遗传育种重点实验室, 北京 100081; 3宁夏农林科学院农作物研究所, 宁夏永宁750105
  • 收稿日期:2016-12-20 修回日期:2017-04-20 出版日期:2017-08-12 网络出版日期:2017-05-11
  • 通讯作者: 郑炜君, E-mail: zhengweijun@nwsuaf.edu.cn; 柴守诚, E-mail: chaishoucheng@126.com
  • 基金资助:

    本研究由国家转基因生物新品种培育重大专项(2014ZX0800916B)和国家自然科学基金项目(31371620)资助。

Soybean Transcription Factor Gene GmNF-YCa Enhances Osmotic Stress Tolerance of Transgenic Arabidopsis

LI Min 1,2,YU Tai-Fei1,2,XU Zhao-Shi2,ZHANG Shuang-Xi 3,MIN Dong-Hong 1,CHEN Ming2,MA You-Zhi2,CHAI Shou-Cheng 1,*,ZHENG Wei-Jun 1,*   

  1. 1 College of Agronomy, Northwest A&F University / State Key Laboratory of Crop Stress Biology for Arid Areas, Yangling 712100, China; 2 Institute of Crop Sciences, Chinese Academy of Agricultural Sciences / National Key Facility for Crop Gene Resources and Genetic Improvement, Key Laboratory of Biology and Genetic Improvement of Triticeae Crops, Ministry of Agriculture, Beijing 100081, China; 3 Institute of Crop Science, Ningxia Academy of Agriculture and Forestry Sciences, Yongning 750105, China
  • Received:2016-12-20 Revised:2017-04-20 Published:2017-08-12 Published online:2017-05-11
  • Contact: zheng Weijun, E-mail: zhengweijun@nwsuaf.edu.cn; Cai Shoucheng, E-mail: chaishoucheng@126.com
  • Supported by:

    This study was supported by the National Major Project for Developing New GM Crops (2014ZX0800916B) and the National Natural Science Foundation of China (31371620).

摘要:

植物的核因子Y(NF-Y)是由3个亚基A、B和C组成,在响应非生物胁迫过程中起着重要的作用。本研究以大豆铁丰8号为材料建立大豆cDNA文库,以pGBKT7-GmDi19-5为诱饵,通过酵母双杂交技术筛选大豆cDNA文库,获得了大豆NF-Y转录因子家族亚基C的一个成员,命名为GmNF-YCa。该基因全长为864 bp,编码287个氨基酸,属于NF-YC亚家族。GmNF-YCa分子量为31.6 kD,等电点为5.07亲水性蛋白,具有一个跨膜结构域,无信号肽。序列分析表明,NF-YC亚家族具有很高的保守性。GmNF-YCa基因启动子含有ARE、Box4、GATA-motif、Box I、ACE、ABRE和CAT-Box等胁迫和光响应元件。组织特异性分析表明,GmNF-YCa基因在种子萌发期表达量最高。实时定量结果表明,GmNF-YCa受蔗糖和甘露醇的诱导上调表达。使用农杆菌介导法将大豆GmNF-YCa基因导入拟南芥,并进行了功能分析。发芽率试验分析表明,GmNF-YCa的转基因提高了转基因拟南芥萌发期对渗透胁迫的耐性;改良了在蔗糖和甘露醇处理下转基因拟南芥的根系生长和侧根发育。

关键词: 大豆, 核因子Y, 表达模式, 启动子, 渗透胁迫

Abstract:

Biotic stresses, like plant diseases and insect pests, and abiotic stresses, such as drought and salt, heavily threaten plant growth and influence crop yield and quality. Osmotic stress resulting from water deficiency in the soil is one of major hurdles. Nuclear Factor Y (NF-Y) is a heterotrimeric protein, consisting of NF-YA, NF-YB and NF-YC in plant. NF-Y plays significant roles in the pathway responding to osmotic stresses in plants. We acquired a gene GmNF-YCa, a member of regulatory subunit NF-YC family in soybean (Glycine max L.), by screening a soybean cDNA library using yeast two-hybrid system. The full sequence of GmNF-YCa is 864 bp, encoding 287 amino acids and having a NF-YC motif, belonging to NF-YC subfamily. GmNF-YCa is a hydrophilic protein with a molecular weight of 31.6 kD and isoelectric point of 5.07, and containing a transmembrane domain and no any signal peptides. Sequence analysis showed that NF-YC subfamily was highly conserved among various species. Promoters of GmNF-YCa contained various abiotic stresses and light responsive elements, such as ARE, Box 4, GATA-motif, Box I, ACE, ABRE,CAT-Box. According to tissue-specific analysis, GmNF-YCa had the highest expression level in germination stage. Quantitative Real-time PCR suggested that GmNF-YCa was induced by sucrose stress and mannitol treatment. GmNF-YCa was transformated to Arabidopsis successfully by Agrobacterim-mediated method and the overexpressed Arabidopsis was prepared for the function characterization analysis. Overexpression of GmNF-YCacould improve tolerance of transgenic Arabidopsis to osmotic stress in the germination stage, and enhance root development with more lateral roots in sucrose and mannitol treatments.

Key words: Soybean, Nuclear Factor Y, Expression pattern, Promoter, Osmotic stress

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