生物技术通报 ›› 2018, Vol. 34 ›› Issue (12): 110-115.doi: 10.13560/j.cnki.biotech.bull.1985.2018-0496

• 研究报告 • 上一篇    下一篇

白桦E-box元件的载体构建及与BplMYB46转录因子的互作分析

王博, 王莲萍, 杨春雨, 国会艳, 魏继承   

  1. 牡丹江师范学院生命科学与技术学院,牡丹江 157011
  • 收稿日期:2018-05-27 出版日期:2018-12-26 发布日期:2018-12-24
  • 作者简介:王博,女,硕士研究生,研究方向:植物分子育种;E-mail:671598750@qq.com
  • 基金资助:
    国家自然科学基金项目(31700587),黑龙江省教育厅备案项目(1352MSYYB002),研究生科技创新项目(kjcx 2018-06mdjnu)

Construction of E-box Motifs and the Analysis of Its Interaction with BplMYB46 Transcription Factor of Betula platyphylla

WANG Bo, WANG Lian-ping, YANG Chun-yu, GUO Hui-yan, WEI Ji-cheng   

  1. Department of Life Science and Technology,Mudanjiang Normal University,Mudanjiang 157011
  • Received:2018-05-27 Published:2018-12-26 Online:2018-12-24

摘要: MYB转录因子是植物中最大的转录因子家族成员之一,主要参与植物次生代谢调控、激素和环境因子的应答,在植物的生长发育中起着至关重要的作用。已有研究发现,E-box的核心序列为CANNTG(N:A/G/C/T),是一类与光响应和苯丙氨酸生物合成途径相关的元件。在前期研究中,先构建顺式作用元件库,然后以转录因子为中心的酵母单杂交技术筛选发现,白桦的BplMYB46转录因子能够与核心序列为CAAATG 的E-box顺式作用元件结合。但是,是否能与E-box的其他核心序列结合还不清楚。本研究将每一种E-box顺式作用元件的核心序列分别进行双链DNA的复性并连接到pHIS2载体上,然后通过酵母单杂交技术筛选与BplMYB46转录因子能够特异结合的E-box顺式作用元件。结果显示,当E-box的核心序列第3个碱基为A/T/C、第四个碱基为A/G/C时,酵母单菌落能在三缺培养基TDO/3AT上生长,表明与BplMYB46转录因子结合的E-box顺式作用元件的特异性序列为CA(A/T/C)(A/G/C)TG。为后续通过BplMYB46转录因子与E-box顺式作用元件的结合来分析BplMYB46转录因子对下游基因的调控,以及为筛选优良下游基因改良白桦的遗传性状奠定数据基础。

关键词: BplMYB46转录因子, E-box顺式作用元件, 酵母单杂交技术

Abstract: MYB transcription factor is one of members in the largest family of transcription factors in plants;it is mainly involved in the regulation of plant secondary metabolism,the response of hormones and environmental factors,and thus plays a crucial role in plant growth and development. A study reveals that the core sequence of E-box is CANNTGN(N:A/G/C/T),which is a cis-element associated with the response to light and phenylpropanoid biosynthesis pathway. A library of cis-acting elements was constructed in our previous study,and results by a transcription factor-centered yeast one-hybrid technology showed that the BplMYB46 transcription factor bound to the E-box cis-acting element with a core sequence of CAAATG in birch;however,it is unclear whether BplMYB46 may bind to other core sequence of E-box. Therefore,in this study,the renaturation of the double-stranded DNA of every core sequence of E-box cis-acting element was performed and connected to the pHIS2 vector. Further,the yeast one-hybrid technique was used to select E-box cis-acting elements that specifically bound to the BplMYB46 transcription factor. The results showed that the yeast colony grew on TDO/3AT medium when the third nucleotide of the core sequence of E-box was A/T/C and the fourth was A/G/C,respectively,indicating that the specific sequence of E-box cis-acting element bound by BplMYB46 transcription factor was CA(A/T/C)(A/G/C)TG. This study provides a data basis for improving the genetic traits of birch by analyzing the regulation of BplMYB46 transcription factor to downstream genes and screening downstream genes via the binding of BplMYB46 transcription factor with E-box cis-acting element,

Key words: BplMYB46 transcription factor, E-box cis-acting element, yeast one-hybrid