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刺葡萄DFR基因克隆及生物信息学分析

赖呈纯 黄贤贵 甘煌灿 潘红 范丽华

赖呈纯, 黄贤贵, 甘煌灿, 潘红, 范丽华. 刺葡萄DFR基因克隆及生物信息学分析[J]. 福建农业学报, 2016, 31(7): 683-689. doi: 10.19303/j.issn.1008-0384.2016.07.003
引用本文: 赖呈纯, 黄贤贵, 甘煌灿, 潘红, 范丽华. 刺葡萄DFR基因克隆及生物信息学分析[J]. 福建农业学报, 2016, 31(7): 683-689. doi: 10.19303/j.issn.1008-0384.2016.07.003
LAI Cheng-chun, HUANG Xian-gui, GAN Huang-can, PAN Hong, FAN Li-Hua. Cloning and Bioinformatics of DFR Gene in Vitis davidii Foёx[J]. Fujian Journal of Agricultural Sciences, 2016, 31(7): 683-689. doi: 10.19303/j.issn.1008-0384.2016.07.003
Citation: LAI Cheng-chun, HUANG Xian-gui, GAN Huang-can, PAN Hong, FAN Li-Hua. Cloning and Bioinformatics of DFR Gene in Vitis davidii Foёx[J]. Fujian Journal of Agricultural Sciences, 2016, 31(7): 683-689. doi: 10.19303/j.issn.1008-0384.2016.07.003

刺葡萄DFR基因克隆及生物信息学分析

doi: 10.19303/j.issn.1008-0384.2016.07.003
基金项目: 

福建省科技计划项目——省属公益类科研院所基本科研专项 2014R1015-6

福建省自然科学基金项目 2016J01126

详细信息
    作者简介:

    赖呈纯(1975-),男,博士,副研究员,主要从事园艺植物生物技术与植物细胞代谢工程研究(E-mail:lccisland@163.com)

    通讯作者:

    范丽华(1957-),女,副研究员,主要从事园艺植物栽培的研究(E-mail:512264119@qq.com)

  • 中图分类号: S663.1;Q785

Cloning and Bioinformatics of DFR Gene in Vitis davidii Foёx

  • 摘要: 根据葡萄DFR基因CDS序列设计刺葡萄开放阅读框(ORF)特异引物,利用RT-PCR技术克隆获得其DFR基因序列,并通过生物信息学方法分析其生物学特性。结果表明,刺葡萄DFR基因ORF序列全长1 014 bp,编码337个氨基酸,命名为Vitis davidii dihydroflavonol 4-reductase gene(VdDFR),GenBank登录号为KF915803。刺葡萄DFR蛋白预测分子量为37 593.2 Da,理论等电点pI为5.81,是一个跨膜亲水蛋白,无典型信号肽,不属于分泌蛋白,并且亚细胞定位主要位于细胞质中(70%);二级结构以无规则卷曲为主(52.82%),是一种mixed类蛋白;该蛋白有潜在的7个糖基化位点和16个磷酸化位点,具有NAD(P)结合位点,有NAD依赖型的表异构酶/脱氢酶的N端结构域,属于NADB_Rossmann超家族成员。核苷酸序列分析表明,刺葡萄DFR基因与美丽葡萄、山葡萄和酿酒葡萄的同源性为99%,与圆叶葡萄同源性为98%,与显齿蛇葡萄同源性为94%,进化上比较保守,利用DFR基因编码区碱基序列所建立的系统关系树与真实的植物进化基本一致。
  • 图  1  DFR基因PCR扩增电泳

    注:M为 Marker DL3000; 1~2为RT-PCR产物。

    Figure  1.  Product of RT-PCR

    图  2  刺葡萄DFR蛋白二级结构GOR4的预测结果

    Figure  2.  Secondary structure of VdDFR protein predicted by GOR4

    图  3  VdDFR蛋白的三级结构模型

    Figure  3.  Tertiary structure of VdDFR protein

    图  4  刺葡萄DFR蛋白氨基酸序列的保守结构域检索

    Figure  4.  Search for conserved domains of amino acid sequence of Vd DFR protein

    图  5  刺葡萄DFR蛋白氨基酸序列的磷酸化位点分析

    Figure  5.  Predicted phosphorylation sites in amino acid sequence of Vd DFR protein

    图  6  植物DFR基因序列构建的系统进化树

    Figure  6.  Phylogenetic tree of plant DFR genes sequences

    表  1  所用的在线分析工具

    Table  1.   Online analytical tools used in this study

    软件来源或网址说明
    ProtParamhttp://web.expasy.org/protparam/蛋白的理化性质分析
    GOR IVhttp://npsa-pbil.ibcp.fr/cgi-bin/npsa_automat.plpage=npsa_gor4.html预测蛋白的二级结构
    PredictProteinhttps://www.predictprotein.org/预测蛋白的二级结构
    Phyre2http://www.sbg.bio.ic.ac.uk/phyre2/html/page.cgiid=indexuk/phyre2/html/page.cgiid=indexuk/phyre2/html/page.cgiid=indexuk/phyre2/html/page.cg预测蛋白的三级结构
    PSORT IIhttp://psort.hgc.jp/form.html蛋白的亚细胞定位
    TargetP 1.1http://www.cbs.dtu.dk/services/TargetP/预测蛋白的信号肽
    TMpredhttp://www.ch.embnet.org/software/TMPRED_form.html蛋白序列的跨膜区分析
    InterProhttp://www.ebi.ac.uk/interpro/scan.html分析预测蛋白保守结构域
    Standard Protein BLASThttp://blast.ncbi.nlm.nih.gov/Blast.cgi?PROGRAM=blastp & PAGE_TYPE=BlastSearch & LINK_LOC=blasthome分析预测蛋白保守结构域
    NetOGlyc 4.0http://www.cbs.dtu.dk/services/NetOGlyc/分析蛋白氨基酸序列潜在的糖基化位点
    NetPhos2.0http://www.cbs.dtu.dk/services/NetPhos/预测蛋白氨基酸序列存在的磷酸化位点
    下载: 导出CSV
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出版历程
  • 收稿日期:  2016-05-17
  • 修回日期:  2016-06-10
  • 刊出日期:  2016-07-01

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