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作物学报 ›› 2011, Vol. 37 ›› Issue (05): 918-923.doi: 10.3724/SP.J.1006.2011.00918

• 研究简报 • 上一篇    下一篇

低温胁迫下东农冬麦1号分蘖节SSH文库的构建及文库中3个基因的表达模式

牟永潮,崔红,于晶,曾俨,孟健男,苍晶*   

  1. 东北农业大学生命科学学院,黑龙江哈尔滨 150030
  • 收稿日期:2010-10-28 修回日期:2011-03-08 出版日期:2011-05-12 网络出版日期:2011-03-24
  • 基金资助:

    本研究由东北农业大学创新团队项目(CXZ003), 东北农业大学科学研究基金和东北农业大学冬小麦创新工程项目资助。

Expression Analysis of Three Genes from SSH Library Constructed Using Tillering Nodes of Dongnongdongmai 1 under Low Temperature

MOU Yong-Chao, CUI Hong, YU Jing, ZENG Yan, MENG Jian-Nan,CANG Jing*   

  1. College of Life Science, Northeast Agricultural University, Harbin 150030, China
  • Received:2010-10-28 Revised:2011-03-08 Published:2011-05-12 Published online:2011-03-24

摘要: 为揭示高抗寒冬小麦品种东农冬麦1号的抗寒分子机制,在北方寒地自然条件下,于越冬前(5℃)和越冬期(-25℃) 分别对分蘖节取样,利用抑制消减杂交技术构建该品种低温胁迫相关基因的cDNA文库。在cDNA文库中随机挑选300个阳性克隆测序,获得230条高质量的表达序列标签(EST)。对该序列进行BLAST比对及功能注释,发现文库中基因组成类型复杂多样,其中应对胁迫的基因(如热激蛋白、CS66Wcor8等)以及参与编码60S和40S核糖体亚基的基因出现的频率较高,这些基因可能与东农冬麦1号的安全越冬有密切关系。对文库中筛选到的2种未知基因和Clp基因进行荧光定量PCR分析,发现其表达水平随着温度降低而不同程度升高;在弱抗寒性小麦品种济麦22中,这些基因则有不同的表达模式。推测这些基因在抗低温胁迫过程中发挥重要作用。

关键词: 东农冬麦1号, 抗寒基因, 抑制消减杂交(SSH), 表达序列标签(EST), 荧光定量PCR

Abstract: To reveal the molecular mechanism of cold resistance of winter wheat variety Dongnongdongmai 1, under natural conditions in the north region of China, we collected tillering nodes before (5°C) and in winter season (-25°C) respectively, then constructed cDNA library of Dongnongdongmai 1 containing low temperature stress related genes by suppression subtractive hybridization. A total of 300 positive clones selected from cDNA library randomly were sequenced to obtain 230 high quality expressed sequence tags (EST). After BLAST and functional annotation, we found complex and diverse types of genes in the library, including stress response genes (such as heat shock protein, CS66, Wcor8 and other related genes) and genes encoding 40S and 60S ribosomal subunit. These genes with high frequency may play an important role in protecting Dongnongdongmai 1 from injuring under low temperature. Two unknown genes and Clp selected from the cDNA library were analyzed through quantitative PCR. The expression levels of these genes in Dongnongdongmai 1 were increased in various degrees with the decrease of temperature; however, their expression patterns were different in Jimai 22 (a winter wheat cultivar with low resistance to low temperature). This result suggests that these genes may play an important role in resistance to low temperature stress.

Key words: Dongnongdongmai 1, Gene for cold resistance, Suppression subtractive hybridization, Expressed sequence tag, Real-time PCR

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