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作物学报 ›› 2009, Vol. 35 ›› Issue (12): 2180-2186.doi: 10.3724/SP.J.1006.2009.02180

• 作物遗传育种·种质资源·分子遗传学 • 上一篇    下一篇

鹰嘴豆锌指蛋白基因ZF1的克隆及表达分析

陈晨1,彭辉1,高文瑞1,石庆华2,张桦2,张巨松2,李建贵2,麻浩1,*   

  1. 1南京农业大学作物遗传与种质创新国家重点实验室/南京农业大学大豆研究所，江苏南京210095；2新疆农业大学农业生物技术研究中心重点实验室，新疆乌鲁木齐830052
  • 收稿日期:2009-02-26 修回日期:2009-07-24 出版日期:2009-12-10 网络出版日期:2009-10-13
  • 通讯作者: 麻浩, E-mail: Lq-ncsi@njau.edu.cn; Tel: 025-84395324
  • 基金资助:

    本研究由国家自然科学基金项目（30860152），国家“十一五”科技支撑计划项目（2007BAC15B06，2006BAD09A04，2006BAD09A08），教育部高等学校学科创新引智计划，“全国科技支疆行动”合作项目（200991254）资助。

Cloning and Expression of Zinc Finger Protein Genetic ZF1 in Chickpea（Cicerone arietinum L.）

CHEN Chen1,PENG Hui1,GAO Wen-Rui1,SHI Qing-Hua2,ZHANG Hua2,ZHANG Ju-Song2,LI Jiang-Gui2,MA Hao1*   

  1. 1State Key Laboratory of Crop Genetics and Germplasm Enhancement/National Center for Soybean Improvement,Nanjing Agricultural University,Nanjing 210095,China;2Key Laboratory of Agricultural Biotechnolgy,Xinjing Agricultural University,Urumqi 830052,China
  • Received:2009-02-26 Revised:2009-07-24 Published:2009-12-10 Published online:2009-10-13
  • Contact: MA Hao,E-mail:Lq-ncsi@njau.edu.cn;Tel:025-84395324

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被引次数

4

摘要：

利用一段从PEG胁迫的鹰嘴豆幼苗叶片所构建的cDNA文库中得到的EST序列，通过3¢RACE方法克隆到一个鹰嘴豆C2H2型锌指蛋白基因ZF1，该基因不含内含子，编码一条244个氨基酸残基的多肽，含有两个典型的Cys2/His2锌指结构。其氨基酸序列含有一个可能的核定位型号，农杆菌介导的洋葱表皮细胞GFP瞬时表达实验表明，ZF1蛋白位于细胞核内。半定量RT-PCR分析表明，ZF1在鹰嘴豆的根、茎、叶、花、幼荚和幼胚中均有表达，在茎和叶中表达较弱，为组成型转录因子。半定量RT-PCR和实时荧光定量PCR检测结果显示，ZF1不但受高温及干旱诱导，而且还受6-苄基腺嘌呤(6-BA)、脱落酸(ABA)、乙烯利(Et)、赤霉素(GA3)、吲哚-3-乙酸(IAA)、茉莉酸甲酯(MeJA)、水杨酸(SA)和氧胁迫诱导。这些结果表明，ZF1基因可能作为一个核调控因子参与植物的生长代谢以及多种生物与非生物胁迫的应答。

关键词: 鹰嘴豆, 锌指蛋白, 基因克隆, 胁迫, 表达分析

Abstract:

Regulation of gene expression at the level of transcription controls many crucial biological processes including growth and development, stress response, signal transduction and disease resistance. A number of factors, such as C2H2 zinc finger protein, are required and factors play an important role in the transcription. Chickpea (Cicer arietinum L.) is the third important legume crop gown mainly in the arid and semi-arid regions in the world. Due to its taxonomic proximity with the model legume genome of Medicago truncatula and its ability to grow in soil with relatively low water content, chickpea is being investigated as a model legume crop for drought tolerance studies. In our laboratory, two cDNA libraries from the PEG-treated and non-treated seedling leaves of chickpea XJ209 were constructed and many genes were found to express differentially and involved in diverse biological processes, such as metabolism, transcription, signal transduction, protein synthesis and others. According to an EST in the cDNA libraries, a zinc finger protein gene ZF1 was cloned by RT-PCR and rapid amplification of cDNA ends (RACE). ZF1 did not include any intron, encoding a 26.33 kD protein with 244 amino acids, containing two typical C2H2 zinc finger domains. The deduced protein sequence had a potential nuclear localization signal (NLS). Meanwhile, transient expression of the ZF1-GFP protein in onion epidermal cells showed that ZF1 protein was localized in cell nuclei. Semi-quantitative RT-PCR analysis showed that ZF1 expressed in root, stem, leaf, flower, immature pod, and embryo of chickpea with different expression patterns. The expression of ZF1 investigated by semi-quantitative PCR had no obvious changes under stresses of cold, salt and wounding, while was increased under the treatments of heat and drought, as well as N-6-benzyl-adenine (6-BA), abscisic acid (ABA), ethephon (Et), gibberellin (GA3), indole-3-acetic acid (IAA), methyl jasmonate (MeJA), salicylic acid (SA), and H2O2. These results from semi-quantitative PCR in several treatments were further confirmed to be mainly in accord with those from real-time quantification PCR. Our results suggest that ZF1 may play multiple roles in abiotic and biotic resistance pathways, as well as in plant growth.

Key words: Chickpea, Zinc finger protein, Gene cloning, Stress, Expression analysis


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