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作物学报 ›› 2017, Vol. 43 ›› Issue (01): 31-41.doi: 10.3724/SP.J.1006.2017.00031

• 作物遗传育种·种质资源·分子遗传学 • 上一篇    下一篇

甘蔗独脚金内酯生物合成关键基因ScD27的克隆与表达分析

吴转娣**,刘新龙,刘家勇,昝逢刚,李旭娟,刘洪博,林秀琴,陈学宽,苏火生,赵培方,吴才文*   

  1. 云南省农业科学院甘蔗研究所 / 云南省甘蔗遗传改良重点实验室,云南开远661699
  • 收稿日期:2016-01-11 修回日期:2016-09-18 出版日期:2017-01-12 网络出版日期:2016-09-28
  • 通讯作者: 吴才文, E-mail: gksky_wcw@163.com
  • 基金资助:

    本研究由国家自然科学基金项目(31360359),国家现代农业产业技术体系建设专项(CARS-20-1-1),云南省中青年学术技术带头人后备人才(2014HB038),云南省应用基础研究计划项目(2016FB071),重大科技专项-生物(2015ZA001)和科技创新人才计划(2014HC015)资助。

Cloning and Expression Analysis of Key Gene ScD27 in Strigolactones Biosynthesis Pathway

WU Zhuan-Di**,LIU Xin-Long**,LIU Jia-Yong,ZAN Feng-Gang,LI Xu-Juan,LIU Hong-Bo,LIN Xiu-Qin, CHEN Xue-Kuan, SU Huo-Sheng,ZHAO Pei-fang,WU Cai-Wen*   

  1. Sugarcane Research Institute, Yunnan Academy of Agricultural Sciences / Yunnan Key Laboratory of Sugarcane Genetic Improvement, Kaiyuan 661699, China
  • Received:2016-01-11 Revised:2016-09-18 Published:2017-01-12 Published online:2016-09-28
  • Contact: 吴才文, E-mail: gksky_wcw@163.com
  • Supported by:

    This study was supported by the National Natural Science Foundation of China (31360359), the National Modern Agricultural Industry Technology System Construction Project (CARS-20-1-1), the Young and Middle-aged Academic Technology Leaders Reserve Talented Person in Yunnan Province (2014HB038), the Applied Basic Research Projects in Yunnan Province (2016FB071), the Major Science and Technology Projects – Biology (2015ZA001), and the Science and Technology Innovation Talents Project (2014HC015).

摘要:

独脚金内酯(Strigolactones SLs)是一类新型植物激素,D27基因是独脚金内酯生物合成途径中最上游的调控基因,且该基因调控SLs的合成是一个可逆的过程。本研究根据水稻、玉米、高粱和二穗短柄草4种禾本科作物的D27基因核苷酸序列保守区域设计引物,以甘蔗品种ROC22的cDNA为模板,利用RT-PCR和RACE技术,从甘蔗中克隆出D27基因的cDNA全长序列,命名为ScD27,GenBank登录号为KP987221.1。该基因序列全长1379 bp,包含一个867 bp的完整开放阅读框(ORF),编码288个氨基酸残基。ScD27编码的蛋白质分子量为71.58 kD,理论等电点为5.04,是一种非分泌性蛋白,主要分布于叶绿体上,该蛋白的保守区可能具有2个锌指蛋白结构域(ZnF_TAZ和ZnF_A20),且不存在信号肽;该基因编码的氨基酸序列具有较高的保守性,与高粱、谷子、大麦和短穗二柄草等禾本科植物的D27氨基酸序列相似性在70%以上;ScD27基因在甘蔗各组织部位均有表达,其中茎尖和腋芽中表达量较高,叶、茎和根中的表达较低。此外,ScD27基因在甘蔗茎尖中的表达受PEG、盐胁迫、磷缺乏和营养缺乏的诱导,推测ScD27基因是甘蔗独脚金内酯生物合成途径中响应非生物胁迫的关键基因。

关键词: 甘蔗, ScD27, 同源克隆, 生物信息学, q-PCR

Abstract:

Strigolactones (SLs) is a novel class of plant hormones. D27 regulating reversible metabolic process is located in up-stream of strigolactones biosynthesis pathway. In this study, primers were designed based on the conserved domains from four species inluding Oryza sativa, Zea mays, Sorghum bicolor, and Brachypodium distachyon. Using cDNA from sugarcane cultivar ROC22 as the template, the full-length cDNA sequence of D27 gene from sugarcane was cloned by RT-PCR and RACE method. This gene isnamed as ScD27, with the GenBank accession number of KP987221.1. Its length is 1379bp, and it contains an 867bp open reading frame (ORF), encoding 288 amino acid residues. ScD27 is not a secretory protein and has a molecular weight of 71.58kD, with a theoretical isoelectric point of 5.04. ScD27 is mainly located in chloroplast and the conserved domains of this protein involve two zinc finger protein structures (ZnF_TAZ and ZnF_A20). Amino acid sequences encoded by ScD27 shared more than 70% similarity with the reported amino acid sequences encoded by D27 of Sorghum bicolor, Setaria italica Beauv., Hordeum vulgare subsp. vulgareand Brachypodium distachyon. ScD27 gene was differentially expressed in different parts of sugarcane plant, with higher level of transcript in stem tip and axillary bud but much lower level in leaf, stem and root. Furthermore, the expression of ScD27 could be induced by the stresses of PEG, salt and the deficiencies of phosphorus and nutrition. These results demonstrated that ScD27 might be a key gene participating in the response to abiotic stresses during sugarcane SLs biosynthesis pathway.

Key words: Sugarcane, ScD27, Homology cloning, Bioinformatics, q-PCR

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