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作物学报 ›› 2017, Vol. 43 ›› Issue (01): 42-50.doi: 10.3724/SP.J.1006.2017.00042

• 作物遗传育种·种质资源·分子遗传学 • 上一篇    下一篇

甘蔗CDK基因的cDNA全长克隆与表达分析

苏亚春,黄珑,凌辉,王竹青,刘峰,苏炜华,黄宁,吴期滨,高世武,阙友雄   

  1. 福建农林大学 / 农业部福建甘蔗生物学与遗传育种重点实验室 / 国家甘蔗产业技术研发中心, 福建福州 350002
  • 收稿日期:2015-12-31 修回日期:2016-07-11 出版日期:2017-01-12 网络出版日期:2016-07-28
  • 通讯作者: 阙友雄, E-mail: queyouxiong@126.com
  • 基金资助:

    本研究由福建省杰出青年科学基金(2015J06006), 福建省高等学校新世纪优秀人才支持计划(JA14095), 引进国际先进农业科学技术计划(948计划)项目(2014-S18)和国家现代农业产业技术体系建设专项(CARS-20)项目资助。

Cloning and Expression Analysis of CDK Gene in Sugarcane

SU Ya-Chun**,HUANG Long**,LING Hui,WANG Zhu-Qing,LIU Feng,SU Wei-Hua,HUANG Ning,WU Qi-Bin, GAO Shi-Wu,QUE You-Xiong*   

  1. Key Laboratory of Sugarcane Biology and Genetic Breeding (Fujian), Ministry of Agriculture / Fujian Agriculture and Forestry University / Sugarcane Research & Development Center, China Agricultural Technology System, Fuzhou 350002, China
  • Received:2015-12-31 Revised:2016-07-11 Published:2017-01-12 Published online:2016-07-28
  • Contact: 阙友雄, E-mail: queyouxiong@126.com
  • Supported by:

    This work was supported by Natural Science Foundation of Fujian Province, China (2015J06006), the Program for New Century Excellent Talents in Fujian Province University (JA14095), the Program of Introducing International Super Agricultural Science and Technology (948 Program) (2014-S18), and the China Agriculture Research System (CARS-20).

摘要:

植物细胞周期依赖性蛋白激酶(cyclin-dependent kinases, CDKs)是一类丝氨酸/苏氨酸蛋白激酶,和周期蛋白(Cyclins)协同作用,是重要的细胞周期性调控因子。本研究从甘蔗受花叶病毒侵染的转录组数据库中获得一条与高粱(Sorghum bicolor) CDK基因(GenBank登录号为XP_002466536.1)高度同源的Unigene序列,通过RT-PCR扩增获得长度为1799 bp的甘蔗ScCDK基因(GenBank登录号为KR258796)。序列分析显示ScCDK基因含一个完整的开放阅读框(65~1603 bp),编码512个氨基酸,且具有CDK典型保守结构域,如ATP结合位点、磷酸结合位点及活化环A-loop。此外,生物信息学预测显示该基因编码的蛋白定位于细胞核,为可溶性蛋白,不存在信号肽,二级结构元件多为无规则卷曲,主要参与蛋白翻译。实时荧光定量PCR分析表明,ScCDK基因的表达具有组织特异性,其在蔗芽上的表达量最高,其次是在蔗肉、蔗根、叶鞘和蔗皮中;在PEG、NaCl和ABA的胁迫诱导过程中,ScCDK均表现上调作用,且受ABA胁迫后表达量最高,约为对照组的1.9倍。推测该基因的表达与甘蔗抗干旱和抗渗透胁迫有关,同时受激素的诱导,参与细胞周期分裂。

关键词: 甘蔗, CDK基因, 同源克隆, 生物信息学分析, 实时荧光定量PCR

Abstract:

Plant cyclin-dependent kinases (CDKs) are a specific class of serine/threonine protein kinases, having synergy with cyclins, as an important acting factor in the process of cell regulation. In this study, a unigene from previous transcriptome data of sugarcane in response to sorghum mosaic virus (SrMV) infection, which exhibited highly homologous to Sorghum biocolor CDK sequence (GenBank Acc No. XP_002466536.1), was validated by RT-PCR and named as ScCDK (GenBank Acc No. KR258796) with a full-length cDNA of 1799 bp. ScCDK contained a complete open reading frame (65–1603 bp) encoding 512 amino acids, and a typical conservative CDK domain, such as ATP binding site, polypeptide substrate binding site and activation loop. Furthermore, based on the bioinformatics prediction, this soluble protein, which was most probably involved in intermediary metabolism, was located in cell nucleus and had no signal peptide but had more random coil upon secondary structure. Real-time quantitative PCR (RT-qPCR) analysis revealed that the expression of this target gene was tissue-specific, with the highest transcript level in bud, followed by the expression in stem pith, root, leaf sheath and epidermis. Besides, its expression could be up-regulated by the treatments of PEG, NaCl and ABA, with nearly 1.9-fold up-regulation of ScCDK transcript under the stress of ABA, compared with the control. These results suggest that the ScCDK may relate to the response of sugarcane to drought and osmotic stresses, meanwhile, be induced by hormones and involved in cell cycle division.

Key words: Sugarcane, CDK, Homology cloning, Bioinformatics analysis, Real-time PCR

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