• 中文核心期刊
  • CSCD来源期刊
  • 中国科技核心期刊
  • CA、CABI、ZR收录期刊

留言板

尊敬的读者、作者、审稿人, 关于本刊的投稿、审稿、编辑和出版的任何问题, 您可以本页添加留言。我们将尽快给您答复。谢谢您的支持!

姓名
邮箱
手机号码
标题
留言内容
验证码

黄肉桃果实八氢番茄红素脱氢酶基因的克隆及表达分析

梁敏华 杨震峰 苏新国 宋春波

梁敏华, 杨震峰, 苏新国, 宋春波. 黄肉桃果实八氢番茄红素脱氢酶基因的克隆及表达分析[J]. 福建农业学报, 2018, 33(6): 608-613. doi: 10.19303/j.issn.1008-0384.2018.06.012
引用本文: 梁敏华, 杨震峰, 苏新国, 宋春波. 黄肉桃果实八氢番茄红素脱氢酶基因的克隆及表达分析[J]. 福建农业学报, 2018, 33(6): 608-613. doi: 10.19303/j.issn.1008-0384.2018.06.012
LIANG Min-hua, YANG Zhen-feng, SU Xin-guo, SONG Chun-bo. Cloning and Expression of Phytoene Desaturase Gene, PpPDS, from Peaches[J]. Fujian Journal of Agricultural Sciences, 2018, 33(6): 608-613. doi: 10.19303/j.issn.1008-0384.2018.06.012
Citation: LIANG Min-hua, YANG Zhen-feng, SU Xin-guo, SONG Chun-bo. Cloning and Expression of Phytoene Desaturase Gene, PpPDS, from Peaches[J]. Fujian Journal of Agricultural Sciences, 2018, 33(6): 608-613. doi: 10.19303/j.issn.1008-0384.2018.06.012

黄肉桃果实八氢番茄红素脱氢酶基因的克隆及表达分析

doi: 10.19303/j.issn.1008-0384.2018.06.012
基金项目: 

国家自然科学基金 31371866

浙江省自然科学基金 LQ15C200004

广东省自然科学基金 2014A03031376

广州市科技计划项目 201804010417

详细信息
    作者简介:

    梁敏华(1987-), 男, 硕士, 主要从事农产品贮藏与加工研究(E-mail: liangmh@gdyzy.edu.cn)

    通讯作者:

    苏新国(1977-), 男, 博士, 教授, 主要从事食品质量与安全研究(E-mail: 31999678@qq.com)

  • 中图分类号: S662.1

Cloning and Expression of Phytoene Desaturase Gene, PpPDS, from Peaches

  • 摘要: 利用基因克隆技术获得桃果实八氢番茄红素脱氢酶基因(PDS)的全长核苷酸序列,命名为PpPDSPpPDS全长2 056 bp,编码区长度1 722 bp,共编码573个氨基酸。进化树分析发现PpPDS与杏果实PaPDS、草莓FaPDS亲缘性较高。蛋白质序列分析表明PpPDS含有二核苷酸[NAD(H)/NADP(H)或FAD(H)]结合基元,其处于NAD(P)-binding Rossmann-like保守结构域内。实时荧光定量PCR结果显示,PpPDS基因在黄肉品种‘金丽’桃果实发育过程中均有表达,且在果肉中的表达普遍高于果皮。果实处于硬核期的PpPDS基因在果实中的表达显著高于软核果时期,随后其表达处于较高水平并呈平缓趋势。本研究对桃果实PpPDS基因的克隆及表达分析为研究桃果实类胡萝卜素生物合成分子机理奠定了良好的基础。
  • 图  1  桃果实PDS基因cDNA PCR扩增

    注:M为1 500 bp DNA Marker;1为RT-PCR产物;2为3'RACE产物;3为5'RACE产物。

    Figure  1.  PCR ampliation of cDNA from peaches for cloning PDS gene

    图  2  PpPDS蛋白功能结构域预测及氨基酸多重序列比对

    注:a为蛋白功能结构域预测;b为基酸序列同源性比较。

    Figure  2.  Prediction of protein functional domains and multiple alignment of putative amino acid sequence on PpPDS gene

    图  3  PpPDS基因系统发生树

    Figure  3.  Phylogenetic trees of PpPDS gene

    图  4  PpPDS蛋白的三维结构预测

    Figure  4.  Deduced 3-dimensional structure of PpPDS protein

    图  5  PpPDS基因在桃果实成熟期间的相对表达量

    注:图中不同小写字母表示在P<0.05水平上差异显著。

    Figure  5.  Relative expression of PpPDS gene during fruit development and maturation of peaches

    表  1  本研究所用引物

    Table  1.   Primers applied

    基因名称 引物序列*[5'-3'] 用途
    PpPDS Forward: CACTGAAGGTTGTTTGCGTNGAYTAYCC
    Reverse: TCCTTAACAGTCAGTCCATCYTGNGCYTC
    中间片段扩增
    3'-GSP1: CCTCGTCCAGCTAAGCCGTTGAAGGTCGTGAT
    3'-GSP2: GGCCATATTGAAGAACAATGAGATGCTGACTT
    3'-RACE PCR扩增
    5'-GSP1: ACAGTCAGTCCATCCTGGGCCTCAACATAAGC
    5'-GSP2: TATCTTTCCATGCTGCCACCTTTCCGCCCAGA
    5'-RACE PCR扩增
    Forward: CCGTTGAAGGTCGTGATTG
    Reverse: CTTTCCGCCCAGAACATC
    实时荧光定量PCR
    PpTEF2 Forward: GGTGTGACGATGAAGAGTGATG;
    Reverse: TGAAGGAGAGGGAAGGTGAAAG
    内参基因
    注:* Y=C/T;N=A/T/C/G。
    下载: 导出CSV
  • [1] 杨勇, 马瑞娟, 沈志军, 等.不同果肉质地桃果实乙烯生成规律及其对乙烯的反应[J].浙江农业学报, 2015, 27(10):1730-1734. doi: 10.3969/j.issn.1004-1524.2015.10.08
    [2] CAPRIOLI I, LAFUENTE M, RODRIGO M, et al. Influence of postharvest treatments on quality, carotenoids, and abscisic acid content of stored 'Spring Belle' peach (Prunus persica) fruit[J]. Journal of Agricultural and Food Chemistry, 2009, 57(15):7056-7063. doi: 10.1021/jf900565g
    [3] CAO S F, LIANG M H, SHI L Y, et al. Accumulation of carotenoids and expression of carotenogenic genes in peach fruit[J].Food Chemistry, 2017, 214:137-146. doi: 10.1016/j.foodchem.2016.07.085
    [4] HUANG J, LIU J, LI Y, et al. Isolation and characterization of the phytoene desaturase gene as a potential selective marker for genetic engineering of the astaxanthin-producing green alga chlorella zofingiensis (chlorophyta)[J]. Journal of Phycology, 2008, 44(3):684-690. doi: 10.1111/jpy.2008.44.issue-3
    [5] 吴磊, 姜朋, 张瑜, 等.苏麦3号小麦穗部病毒诱导的基因沉默(VIGS)体系的建立及验证[J].江苏农业学报, 2017, 33(2):248-252. http://www.cqvip.com/QK/92499X/201702/WD748378898866504849554850484851.html
    [6] VAN DIEN S J, MARX C J, O'BRIEN B N, et al. genetic characterization of the carotenoid biosynthetic pathway in methylobacterium extorquens AM1 and isolation of a colorless mutant[J]. Applied and Environmental Microbiology, 2003, 69(12):7563-7566. doi: 10.1128/AEM.69.12.7563-7566.2003
    [7] CHEN C, BAI L H, QIAO D R, et al. Cloning and expression study of a putative carotene biosynthesis related (cbr) gene from the halotolerant green alga Dunaliella salina[J]. Molecular Biology Reports, 2008, 35(3):321-327. doi: 10.1007/s11033-007-9089-z
    [8] FURUBAYASHI M, SAITO K, UMENO D. Evolutionary analysis of the functional plasticity of Staphylococcus aureus C30 carotenoid synthase[J]. Journal of Bioscience & Bioengineering, 2014, 117(4):431-436. http://ci.nii.ac.jp/naid/110009817057
    [9] GR NEWALD K, ECKERT M, HIRSCHBERG J, et al. Phytoene desaturase is localized exclusively in the chloroplast and up-regulated at the mRNA level during accumulation of secondary carotenoids in Haematococcus pluvialis (Volvocales, chlorophyceae)[J]. Plant Physiology, 2000, 122(4):1261-1268. doi: 10.1104/pp.122.4.1261
    [10] RS A, FE D, A M, et al. Characterization of a higher plant herbicide-resistant phytoene desaturase and its use as a selectable marker[J].Plant Biotechnology Journal, 2006, 4(2):263-273. doi: 10.1111/pbi.2006.4.issue-2
    [11] MARTY I, BUREAU S, SARKISSIAN G, et al. Ethylene regulation of carotenoid accumulation and carotenogenic gene expression in colour-contrasted apricot varieties (Prunus armeniaca)[J].Journal of Experimental Botany, 2005, 56(417):1877-1886. doi: 10.1093/jxb/eri177
    [12] YAN P, GAO X Z, SHEN W T, et al. Cloning and expression analysis of phytoene desaturase and ζ-carotene desaturase genes in Carica papaya[J]. Molecular Biology Reports, 2011, 38(2):785-791. doi: 10.1007/s11033-010-0167-2
    [13] WISUTIAMONKUL A, AMPOMAH-DWAMENA C, ALLAN A C, et al. Carotenoid accumulation and gene expression during durian (Durio zibethinus) fruit growth and ripening[J]. Scientia Horticulturae, 2017, 220:233-242. doi: 10.1016/j.scienta.2017.03.048
    [14] TONG Z, GAO Z, WANG F, et al. Selection of reliable reference genes for gene expression studies in peach using real-time PCR[J]. BMC Molecular Biology, 2009, 10(1):71-83. doi: 10.1186/1471-2199-10-71
    [15] FRIGAARD N U, MARESCA J A, YUNKER C E, et al. Genetic manipulation of carotenoid biosynthesis in the green sulfur bacterium Chlorobium tepidum[J].Journal of Bacteriology, 2004, 186(16):5210-5220. doi: 10.1128/JB.186.16.5210-5220.2004
    [16] 吴建设, 钟淮钦, 黄敏玲.观赏向日葵八氢番茄红素脱氢酶基因PDS的cDNA克隆与表达特性分析[J].中国细胞生物学学报, 2013, 35(12):1753-1759. doi: 10.11844/cjcb.2013.12.0234
    [17] BLAS A L, MING R, LIU Z, et al. Cloning of the papaya chromoplast-specific lycopene beta-cyclase, CpCYC-b, controlling fruit flesh color reveals conserved microsynteny and a recombination hot spot[J]. Plant Physiology, 2010, 152(4):2013-2022. doi: 10.1104/pp.109.152298
    [18] BRAMLEY P M. Regulation of carotenoid formation during tomato fruit ripening and development[J].Journal of Experimental Botany, 2002, 53(377):2107-2113. doi: 10.1093/jxb/erf059
    [19] CUNNINGHAM F X. Regulation of carotenoid synthesis and accumulation in plants[J].Pure and Applied Chemistry, 2002, 74(8):1409-1417. doi: 10.1351/pac200274081409
    [20] QIN G, GU H, MA L, et al. Disruption of phytoene desaturase gene results in albino and dwarf phenotypes in Arabidopsis by impairing chlorophyll, carotenoid, and gibberellin biosynthesis[J]. Cell Research, 2007, 17(5):471-482. doi: 10.1038/cr.2007.40
  • 加载中
图(5) / 表(1)
计量
  • 文章访问数:  999
  • HTML全文浏览量:  179
  • PDF下载量:  14
  • 被引次数: 0
出版历程
  • 收稿日期:  2018-01-12
  • 修回日期:  2018-04-12
  • 刊出日期:  2018-06-01

目录

    /

    返回文章
    返回