• 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

木薯MeERF1.2基因克隆及表达分析

曾坚 吴伟姗 谢彩虹 沈梓欣 叶晓雪 颜彦 李冰 陈志晟 彭红元 胡伟 曾力旺

曾坚,吴伟姗,谢彩虹,等. 木薯MeERF1.2基因克隆及表达分析 [J]. 福建农业学报,2023,38(4):410−416 doi: 10.19303/j.issn.1008-0384.2023.04.003
引用本文: 曾坚,吴伟姗,谢彩虹,等. 木薯MeERF1.2基因克隆及表达分析 [J]. 福建农业学报,2023,38(4):410−416 doi: 10.19303/j.issn.1008-0384.2023.04.003
ZENG J, WU W S, XIE C H, et al. Cloning and Expression of MeERF1.2 in Cassava [J]. Fujian Journal of Agricultural Sciences,2023,38(4):410−416 doi: 10.19303/j.issn.1008-0384.2023.04.003
Citation: ZENG J, WU W S, XIE C H, et al. Cloning and Expression of MeERF1.2 in Cassava [J]. Fujian Journal of Agricultural Sciences,2023,38(4):410−416 doi: 10.19303/j.issn.1008-0384.2023.04.003

木薯MeERF1.2基因克隆及表达分析

doi: 10.19303/j.issn.1008-0384.2023.04.003
基金项目: 广东省基础与应用基础研究基金项目(2021A1515011236、 2023A1515010336);中央级公益性科研院所基本科研业务费专项(1630052022027);海南省自然科学基金面上项目(320MS098、 322MS128);韶关学院重点项目(SZ2022KJ05);广东省普通高校重点领域专项(2022ZDZX4047);国家自然科学基金项目(31901537);广东省教育厅重大项目(2017KZDXM077);校级大学生创新创业训练计划项目(Sycxcy2022089)
详细信息
    作者简介:

    曾坚 (1987−),男,博士,副教授,研究方向:植物基因功能研究(E-mail:zengjian@sgu.edu.cn

    通讯作者:

    曾力旺(1987−),女,硕士,助理研究员,研究方向:植物基因分子生物学(E-mail:zengliwang@163.com

  • 中图分类号: S533

Cloning and Expression of MeERF1.2 in Cassava

  • 摘要:   目的  乙烯响应因子(Ethylene response factor,ERF)是乙烯信号转导通路的重要成员,克隆并分析其在木薯块根采后生理性变质(Post-harvest physiological deterioration,PPD)过程中的表达情况,能为进一步研究乙烯信号在木薯PPD过程中的功能提供参考。  方法  以木薯栽培品种华南8号(SC8)为材料,采用RT-PCR技术克隆木薯MeERF1.2基因,对其进行相关生物信息学分析,如遗传进化关系、结构域、蛋白质结构预测、理化性质等。对MeERF1.2基因在细胞中的亚细胞定位进行确认,并用qRT-PCR技术分析MeERF1.2基因在木薯块根PPD过程中的表达水平。  结果  克隆得到的MeERF1.2基因全长为660 bp,编码的氨基酸残基数为219,分子量和等电点分别为25.04 kD和5.61,含有AP2家族结构域,和橡胶HbERF1B-like的亲缘关系最近,序列相似性达到88.74%。MeERF1.2基因定位于细胞核。和对照0 h相比,MeERF1.2基因的表达量在木薯块根的采后过程中表现为显著上升趋势,即MeERF1.2基因的表达受到PPD过程的诱导。  结论  克隆得到的MeERF1.2基因包含ERF基因家族的保守结构域,属于ERF基因家族;MeERF1.2基因的表达在采后过程中受到诱导,可能参与了木薯块根的PPD过程,为后续进一步分析乙烯信号转导通路在PPD过程中的作用奠定了基础。
  • 图  1  MeERF1.2基因扩增结果

    M:DNA Marker;1:样品; 2:阴性对照。

    Figure  1.  PCR amplification of MeERF1.2

    M: DNA Marker; 1: sample; 2: negative control.

    图  2  结构域分析

    Figure  2.  Analysis of conserved domain

    图  3  MeERF1.2蛋白序列的同源性比对

    Figure  3.  Homologous alignment of MeERF1.2 sequences

    图  4  ERF基因的系统进化树

    Figure  4.  Phylogenetic tree of ERF genes

    图  5  MeERF1.2蛋白亚细胞定位分析

    Figure  5.  Subcellular localization of MeERF1.2 protein

    图  6  MeERF1.2在不同组织中的表达量

    L:叶,M:中脉,S:茎,P:叶柄,LB:侧芽,OES:分化胚组织,FR:须根,SR:根,RAM:根顶端分生组织。不同小写字母表明在Duncan’s多重比较中差异显著(P<0.05)

    Figure  6.  Expressions of MeERF1.2 in tissues

    L: leaf; M: midvein; S: stem; P: petiole; LB: lateral bud; OES: organized embryogenic structure; FR: fibrous root; SR: storage root; RAM: root apical meristem. Data with different letters indicate significant differences based on Duncan's multiple range tests (P<0.05).

    图  7  MeERF1.2基因在PPD中的表达变化

    A MeERF1.2基因在采后腐烂过程中转录组测序表达量;B: MeERF1.2基因在采后腐烂过程中实时定量PCR检测结果;*和**分别代表与0 h差异显著(P<0.05)和极显著(P<0.01)。

    Figure  7.  Expressions of MeERF1.2 during PPD

    A: expression of MeERF1.2 during cassava PPD shown by transcriptome sequencing; B: expression of MeERF1.2 during cassava PPD shown by qRT-PCR; * and ** represent significantly different from control (0 h) at P<0.05 and P<0.01, respectively.

    表  1  扩增引物序列

    Table  1.   Primers and their sequences

    引物名称
    Primer
    引物序列
    Primer sequence
    引物用途
    Primer usage
    MeERF1.2-F5′-ATGGATTCCTCCATCTTTCATTCT-3′基因克隆
    MeERF1.2-R5′-CCAAGGTCTTATAGCATTCTCAGAT-3′
    BiMeERF1.2-F5′-AGTGGTCTCTGTCCAGTCCTATGGATTCCTCCATCTTT-3′亚细胞定位
    BiMeERF1.2-R5′-GGTCTCAGCAGACCACAAGTCCAAGGTCTTATAGCATT-3′
    qMeERF1.2-F5′-GAGCTGGGGCTGTACTCAAT-3′荧光定量PCR
    qMeERF1.2-R5′-CAGGTGAGCACCCTTCTTCT-3′
    MeEF1-F5′-TGAACCACCCTGGTCAGATTGGAA-3′内参基因
    MeEF1-R5′-AACTTGGGCTCCTTCTCAAGCTCT-3′
    下载: 导出CSV
  • [1] 张鹏, 杨俊, 周文智, 等. 能源木薯高淀粉抗逆分子育种研究进展与展望 [J]. 生命科学, 2014, 26(5):465−473.

    ZHANG P, YANG J, ZHOU W Z, et al. Progress and perspective of cassava molecular breeding for bioenergy development [J]. Chinese Bulletin of Life Sciences, 2014, 26(5): 465−473.(in Chinese)
    [2] HU W, KONG H, GUO Y L, et al. Comparative physiological and transcriptomic analyses reveal the actions of melatonin in the delay of postharvest physiological deterioration of cassava [J]. Frontiers in Plant Science, 2016, 7: 736.
    [3] 郭靖, 章玉香, 黄芷颐, 等. 木薯MePYL12基因克隆及采后生理性变质过程的表达分析 [J]. 福建农业学报, 2021, 36(1):17−23.

    GUO J, ZHANG Y X, HUANG Z Y, et al. Cloning and post-harvest physiological deterioration expression of MePYL12 of cassava [J]. Fujian Journal of Agricultural Sciences, 2021, 36(1): 17−23.(in Chinese)
    [4] WESTBY A. Cassava utilization, storage and small-scale processing[M]//Cassava: biology, production and utilization. UK: CABI Publishing, 2001: 281-300.
    [5] 马秋香, 许佳, 乔爱民, 等. 木薯储藏根采后生理性变质研究进展 [J]. 热带亚热带植物学报, 2009, 17(3):309−314.

    MA Q X, XU J, QIAO A M, et al. Current progress in studies on post-harvest physiological deterioration of cassava storage roots [J]. Journal of Tropical and Subtropical Botany, 2009, 17(3): 309−314.(in Chinese)
    [6] IYER S, MATTINSON D S, FELLMAN J K. Study of the early events leading to cassava root postharvest deterioration [J]. Tropical Plant Biology, 2010, 3(3): 151−165. doi: 10.1007/s12042-010-9052-3
    [7] HERSHKOVITZ V, FRIEDMAN H, GOLDSCHMIDT E E, et al. Induction of ethylene in avocado fruit in response to chilling stress on tree [J]. Journal of Plant Physiology, 2009, 166(17): 1855−1862. doi: 10.1016/j.jplph.2009.05.012
    [8] 赵赫, 陈受宜, 张劲松. 乙烯信号转导与植物非生物胁迫反应调控研究进展 [J]. 生物技术通报, 2016, 32(10):1−10.

    ZHAO H, CHEN S Y, ZHANG J S. Ethylene signaling pathway in regulating plant response to abiotic stress [J]. Biotechnology Bulletin, 2016, 32(10): 1−10.(in Chinese)
    [9] 施怡婷, 杨淑华. 中国科学家在乙烯信号转导领域取得突破性进展 [J]. 植物学报, 2016, 51(3):287−289.

    SHI Y T, YANG S H. Chinese scientists made breakthrough in study on ethylene signaling transduction in plants [J]. Chinese Bulletin of Botany, 2016, 51(3): 287−289.(in Chinese)
    [10] REN M Y, FENG R J, SHI H R, et al. Expression patterns of members of the ethylene signaling-related gene families in response to dehydration stresses in cassava [J]. PLoS One, 2017, 12(5): e0177621. doi: 10.1371/journal.pone.0177621
    [11] CAsO W H, LIU J, CHEN T, et al. Ethylene receptor signaling and plant salt-stress responses[C]//RAMINA A, CHANG C, GIOVANNONI J, et al. Advances in Plant Ethylene Research. Dordrecht: Springer, 2007: 333-339.
    [12] 于延文, 黄荣峰. 乙烯与植物抗逆性 [J]. 中国农业科技导报, 2013, 15(2):70−75.

    YU Y W, HUANG R F. Ethylene and plant resistance to adversity [J]. Journal of Agricultural Science and Technology, 2013, 15(2): 70−75.(in Chinese)
    [13] DESIKAN R, LAST K, HARRETT-WILLIAMS R, et al. Ethylene-induced stomatal closure in Arabidopsis occurs via AtrbohF-mediated hydrogen peroxide synthesis [J]. The Plant Journal:for Cell and Molecular Biology, 2006, 47(6): 907−916. doi: 10.1111/j.1365-313X.2006.02842.x
    [14] 杨洋. 拟南芥乙烯合成因子依赖ROS代谢调节盐胁迫反应[D]. 开封: 河南大学, 2011.

    YANG Y. The ethylene biosynthesis factors mediate Arabidopsis tolerance to NaCl stress in dependence on ROS metabolism[D]. Kaifeng: Henan University, 2011. (in Chinese)
    [15] MA Y R, YANG M N, WANG J J, et al. Application of exogenous ethylene inhibits postharvest peel browning of ‘Huangguan’ pear [J]. Frontiers in Plant Science, 2017, 7: 2029.
    [16] BLEECKER A B, KENDE H. Ethylene: A gaseous signal molecule in plants [J]. Annual Review of Cell and Developmental Biology, 2000, 16: 1−18. doi: 10.1146/annurev.cellbio.16.1.1
    [17] ALONSO J M, STEPANOVA A N, SOLANO R, et al. Five components of the ethylene-response pathway identified in a screen for weak ethylene-insensitive mutants in Arabidopsis [J]. Proceedings of the National Academy of Sciences of the United States of America, 2003, 100(5): 2992−2997. doi: 10.1073/pnas.0438070100
    [18] ZHAO Q, GUO H W. Paradigms and paradox in the ethylene signaling pathway and interaction network [J]. Molecular Plant, 2011, 4(4): 626−634. doi: 10.1093/mp/ssr042
    [19] 史庆玲, 李忠峰, 董永彬, 等. 植物乙烯信号转导通路及其相关基因的研究进展 [J]. 生物技术进展, 2019, 9(5):449−454.

    SHI Q L, LI Z F, DONG Y B, et al. Progress on ethylene signal transduction pathway and related genes in plants [J]. Current Biotechnology, 2019, 9(5): 449−454.(in Chinese)
    [20] 葛宝宇, 林轶, 侯和胜. ERF类转录因子的结构与功能 [J]. 安徽农学通报, 2007, 13(20):32−35. doi: 10.3969/j.issn.1007-7731.2007.20.013

    GE B Y, LIN Y, HOU H S. Structure and function of ERF transcription factors [J]. Anhui Agricultural Science Bulletin, 2007, 13(20): 32−35.(in Chinese) doi: 10.3969/j.issn.1007-7731.2007.20.013
    [21] NAKANO T, SUZUKI K, FUJIMURA T, et al. Genome-wide analysis of the ERF gene family in Arabidopsis and rice [J]. Plant Physiology, 2006, 140(2): 411−432. doi: 10.1104/pp.105.073783
    [22] SHOJI T, MISHIMA M, HASHIMOTO T. Divergent dna-binding specificities of a group of ethylene response factor transcription factors involved in plant defense [J]. Plant Physiology, 2013, 162(2): 977−990. doi: 10.1104/pp.113.217455
    [23] ABIRI R, SHAHARUDDIN N A, MAZIAH M, et al. Role of ethylene and the APETALA 2/ethylene response factor superfamily in rice under various abiotic and biotic stress conditions [J]. Environmental and Experimental Botany, 2017, 134: 33−44. doi: 10.1016/j.envexpbot.2016.10.015
    [24] SOLANO R, STEPANOVA A, CHAO Q, et al. Nuclear events in ethylene signaling: A transcriptional cascade mediated by ETHYLENE-INSENSITIVE3 and ETHYLENE-RESPONSE-FACTOR1 [J]. Genes & Development, 1998, 12(23): 3703−3714.
    [25] QIN F, SAKUMA Y, LI J, et al. Cloning and functional analysis of a novel DREB1/CBF transcription factor involved in cold-responsive gene expression in Zea mays L [J]. Plant and Cell Physiology, 2004, 45(8): 1042−1052. doi: 10.1093/pcp/pch118
    [26] VANDERSCHUREN H, NYABOGA E, POON J S, et al. Large-scale proteomics of the cassava storage root and identification of a target gene to reduce postharvest deterioration [J]. The Plant Cell, 2014, 26(5): 1913−1924. doi: 10.1105/tpc.114.123927
    [27] 张云川, 曹天澳, 雷骥良, 等. 橡胶草ERF3基因的克隆、亚细胞定位及表达分析 [J]. 植物生理学报, 2021, 57(6):1261−1270.

    ZHANG Y C, CAO T A, LEI J L, et al. Cloning, subcellular localization and expression analysis of TkERF3 in Taraxacum kok-saghyz [J]. Plant Physiology Journal, 2021, 57(6): 1261−1270.(in Chinese)
    [28] JUNG H, CHUNG P J, PARK S H, et al. Overexpression of OsERF48 causes regulation of OsCML16, a calmodulin-like protein gene that enhances root growth and drought tolerance [J]. Plant Biotechnology Journal, 2017, 15(10): 1295−1308. doi: 10.1111/pbi.12716
    [29] LEE D K, YOON S, KIM Y S, et al. Rice OsERF71-mediated root modification affects shoot drought tolerance [J]. Plant Signaling & Behavior, 2017, 12(1): e1268311.
    [30] ZHANG G Y, CHEN M, LI L C, et al. Overexpression of the soybean GmERF3 gene, an AP2/ERF type transcription factor for increased tolerances to salt, drought, and diseases in transgenic tobacco [J]. Journal of Experimental Botany, 2009, 60(13): 3781−3796. doi: 10.1093/jxb/erp214
    [31] CHARFEDDINE M, CHARFEDDINE S, GHAZALA I, et al. Investigation of the response to salinity of transgenic potato plants overexpressing the transcription factor StERF94 [J]. Journal of Biosciences, 2019, 44(6): 141. doi: 10.1007/s12038-019-9959-2
    [32] SHIN S Y, PARK M H, CHOI J W, et al. Gene network underlying the response of harvested pepper to chilling stress [J]. Journal of Plant Physiology, 2017, 219: 112−122. doi: 10.1016/j.jplph.2017.10.002
  • 加载中
图(7) / 表(1)
计量
  • 文章访问数:  396
  • HTML全文浏览量:  141
  • PDF下载量:  18
  • 被引次数: 0
出版历程
  • 收稿日期:  2022-06-09
  • 录用日期:  2022-06-09
  • 修回日期:  2022-11-15
  • 网络出版日期:  2023-03-28
  • 刊出日期:  2023-04-28

目录

    /

    返回文章
    返回