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园艺学报 ›› 2019, Vol. 46 ›› Issue (6): 1033-1044.doi: 10.16420/j.issn.0513-353x.2018-0739

• 研究论文 • 上一篇    下一篇

苹果乙烯响应因子MdERF1B-like的克隆与功能鉴定

张 静1,慈志娟2,许海峰1,姜生辉1,房鸿成1,王意程1,张宗营1,杨官显1,陈学森1,*   

  1. 1山东农业大学园艺科学与工程学院/作物生物学国家重点实验室,山东泰安 271018;2山东省烟台市农业科学研究院,山东烟台 265500
  • 出版日期:2019-06-25 发布日期:2019-06-25
  • 基金资助:
    国家重点研发计划项目(2016YFC0501505);国家自然科学基金项目(31730080,31572091)

Cloning and Functional Characterization of an Ethylene Response Factor MdERF1B-like in Apple

ZHANG Jing1,CI Zhijuan2,XU Haifeng1,JIANG Shenghui1,FANG Hongcheng1,WANG Yicheng1,ZHANG Zongying1,YANG Guanxian1,and CHEN Xuesen1,*   

  1. 1College of Horticulture Science and Engineering,Shandong Agricultural University,State Key Laboratory of Crop Biology,Tai’an,Shandong 271018,China;2Yantai Academy of Agricultural Sciences,Yantai,Shandong 265500,China
  • Online:2019-06-25 Published:2019-06-25

摘要: 以‘泰山早霞’苹果发育期中未着色和着色的果实为试材,利用qRT-PCR分析花青苷和乙烯通路相关基因的表达。分离出1个乙烯响应因子,暂命名为MdERF1B-like(MDP0000167207),其开放阅读框为690 bp,编码229个氨基酸。在‘泰山早霞’苹果着色与未着色果实中,MdERF1B-like表达量变化与花青苷合成基因MdDFR、MdANS和调控基因MdMYB9的表达趋势一致。系统进化树分析表明,MdERF1B-like位于第Ⅸ组,与PyERF1B-like亲缘关系最近。在‘王林’苹果愈伤组织中过表达MdERF1B-like,其花青苷含量显著高于对照。分析MdMYB9启动子序列,其长度为746 bp,序列中含有1个ERF潜在结合元件RAA(TGTTG),酵母单杂表明MdERF1B-like与MdMYB9启动子结合。进一步通过荧光素酶报告试验验证MdERF1B-like促进MdMYB9启动子的转录活性。因此,MdERF1B-like可能通过对MdMYB9的调控来促进苹果花青苷积累。

关键词: 苹果, 乙烯响应因子, 酵母单杂, 荧光素酶报告, 花青苷

Abstract: The expression levels of genes involved in anthocyanin and ethylene biosythesis were analyzed via qRT-PCR using uncolored and colored fruits of‘Taishan Zaoxia’apple at different developmental stages. An ethylene response factor was isolated and tentatively named as MdERF1B-like (MDP0000167207),with an open reading frame of 690 bp encoding a protein of 229 amino acid residues. The expression level of MdERF1B-like was consistent with the expression trends of anthocyanin synthesis related genes MdDFR,MdANS and the regulatory gene MdMYB9 in the colored and uncolored‘Taishan Zaoxia’fruit. Phylogenic analysis indicated that MdERF1B-like was located in group IX and was closely related to PyERF1B-like. Overexpression of MdERF1B-like in‘Orin’calli led to the significantly higher anthocyanin accumulation than in the control. The MdMYB9 promoter sequence of 746 bp was analyzed,which contained an ERF binding RAA motif in its promoter,and yeast one-hybrid assay showed that MdERF1B-like bound with MdMYB9 promoter. Furthermore,luciferase reporter assay was used to verify that MdERF1B-like can promote the transcriptional activity of MdMYB9 promoter. Therefore,MdERF1B-like may promote the accumulation of anthocyanin through regulating the expression of MdMYB9.

Key words: apple, ethylene response factor, yeast one-hybrid assay, luciferase reporter assay, anthocyanin

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