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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
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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

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