中国农业科学 ›› 2022, Vol. 55 ›› Issue (10): 1877-1890.doi: 10.3864/j.issn.0578-1752.2022.10.001
刘瑞达1,2(),葛常伟2,王敏轩1,2,申延会2,李朋珍2,崔子倩2,刘瑞华2,沈倩2,张思平2,刘绍东2,马慧娟2,陈静2,张桂寅1(
),庞朝友2,3(
)
收稿日期:
2021-12-22
接受日期:
2022-01-28
出版日期:
2022-05-16
发布日期:
2022-06-02
通讯作者:
张桂寅,庞朝友
作者简介:
刘瑞达,E-mail: 基金资助:
LIU RuiDa1,2(),GE ChangWei2,WANG MinXuan1,2,SHEN YanHui2,LI PengZhen2,CUI ZiQian2,LIU RuiHua2,SHEN Qian2,ZHANG SiPing2,LIU ShaoDong2,MA HuiJuan2,CHEN Jing2,ZHANG GuiYin1(
),PANG ChaoYou2,3(
)
Received:
2021-12-22
Accepted:
2022-01-28
Online:
2022-05-16
Published:
2022-06-02
Contact:
GuiYin ZHANG,ChaoYou PANG
摘要:
【目的】MYB基因家族作为植物中最大的转录因子家族之一,在抵御逆境胁迫中发挥着重要的作用。克隆陆地棉MYB转录因子基因GhMYB108,并进行表达分析,验证其在干旱胁迫响应中的作用,为进一步研究GhMYB108调控陆地棉耐旱的分子机制奠定基础。【方法】根据干旱转录组数据分析,确定GhMYB108为干旱响应基因;运用聚合酶链式反应(PCR)从陆地棉根系cDNA中扩增目的基因;对GhMYB108进行基因结构特征、预测基因序列信息以及系统进化关系等生物信息学分析;利用Plant Care网站对获得的基因启动子序列进行分析;在不同逆境胁迫条件下,对GhMYB108的表达特性进行qRT-PCR分析;通过亚细胞定位确定GhMYB108蛋白在细胞中的位置;利用酵母试验验证其转录活性;使用病毒诱导的基因沉默技术(virus induced gene silencing,VIGS)沉默GhMYB108,并用qRT-PCR检测基因沉默效率。观察沉默株系在干旱处理前后的表型变化,并统计存活率,采用试剂盒测定相关生理生化指标;通过对棉花叶片喷施ABA与氟啶酮试验来分析GhMYB108与ABA的关系。【结果】从陆地棉中克隆了GhMYB108(Gh_A10G1563),其全长879 bp,编码292个氨基酸,其蛋白质相对分子量为33.288 kD,等电点为6.037,多重序列比对和保守结构域分析,发现GhMYB108含有2个高度保守的MYB结合结构域,属于典型的R2R3型MYB转录因子。不同物种亲缘关系分析发现,GhMYB108与AtMYB108、AtMYB78和AtMYB2的同源性较高,属于同一亚族,且已有研究发现AtMYB108、AtMYB78和AtMYB2与干旱或ABA信号通路相关。GhMYB108定位于细胞核,且具有转录激活活性。在干旱和对照植株中,GhMYB108均在根中表达量最高,茎中表达量最低,并且受自然干旱、18% PEG 6000模拟干旱、盐胁迫和低温等非生物胁迫诱导表达。GhMYB108沉默之后,在自然干旱条件下,沉默植株出现临界表型,与对照相比,其萎蔫更严重,且存活率降低,一些生理生化指标也发生显著变化,如叶片失水率加快,丙二醛含量升高,叶片相对含水量和脯氨酸含量减少,过氧化氢酶(CAT)和过氧化物酶(POD)活性降低,且通过DAB与NBT染色发现植物体积累了更多过氧化氢(H2O2)和超氧阴离子(O2-)。通过对棉花叶片喷施激素ABA或氟啶酮发现GhMYB108可受ABA信号的正调控。【结论】GhMYB108正调控棉花抗旱性,且受ABA信号的正调控。
刘瑞达, 葛常伟, 王敏轩, 申延会, 李朋珍, 崔子倩, 刘瑞华, 沈倩, 张思平, 刘绍东, 马慧娟, 陈静, 张桂寅, 庞朝友. 陆地棉转录因子基因GhMYB108的克隆及其在抗旱中的作用[J]. 中国农业科学, 2022, 55(10): 1877-1890.
LIU RuiDa, GE ChangWei, WANG MinXuan, SHEN YanHui, LI PengZhen, CUI ZiQian, LIU RuiHua, SHEN Qian, ZHANG SiPing, LIU ShaoDong, MA HuiJuan, CHEN Jing, ZHANG GuiYin, PANG ChaoYou. Cloning and Drought Resistance Analysis of Transcription Factor GhMYB108 in Gossypium hirsutum [J]. Scientia Agricultura Sinica, 2022, 55(10): 1877-1890.
表1
GhMYB108启动子序列中抗逆和激素相关顺式作用元件"
位点名称 Site name | 序列 Sequence | 位点功能 Function of site |
---|---|---|
G-Box | CACGTT | 参与光反应的顺式调节元件Cis regulatory elements involved in photoreaction |
ABRE | ACGTG | 参与脱落酸反应的顺式作用元件Cis acting elements involved in abscisic acid reaction |
CGTCA-motif | CGTCA | 参与MeJA响应的顺式调控元件Cis regulatory elements involved in MeJA response |
TC-rich repeats | GTTTTCTTAC | 参与防御和应激反应的顺式作用元件Cis acting elements involved in defense and stress response |
TGA-element | AACGAC | 生长素响应顺式调控元件Auxin responsive Cis regulatory element |
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