芒果MiZAT10A和MiZAT10B功能分析

doi:10.3864/j.issn.0578-1752.2023.01.015

摘要/Abstract

摘要：

【目的】锌指蛋白（zinc finger protein，ZFP）在植物非生物胁迫应答中起重要的作用，研究两个锌指蛋白基因MiZAT10A和MiZAT10B转入拟南芥对盐、干旱、重金属以及外源激素等非生物胁迫的应答，为抗逆育种提供理论依据。【方法】利用在线软件PLACE和MEME分别对芒果MiZAT10A和MiZAT10B进行启动子顺式作用元件以及motif预测和分析，并利用TBtools软件和‘四季蜜芒’基因注释文件（GFF文件，未公开）绘制染色体定位图；通过实时荧光定量分析MiZAT10A和MiZAT10B的组织表达模式；构建芒果MiZAT10A和MiZAT10B超量表达载体，采用农杆菌花序浸染法转化模式植物拟南芥，观察并记录转基因拟南芥开花表型以及在盐、干旱、重金属以及外源激素脱落酸和赤霉素处理下的根生长情况。【结果】启动子顺式元件分析显示，两个基因的启动子区域都有许多光响应元件、激素响应元件和非生物胁迫响应元件。表达模式分析显示，MiZAT10A与MiZAT10B在芽和花中表达水平最高。MiZAT10A和MiZAT10B分别获得了9株和14株转基因拟南芥，开花表型分析显示，MiZAT10A和MiZAT10B转基因拟南芥提早开花。在盐胁迫、干旱胁迫和重金属胁迫以及GA₃和ABA激素处理下，两个超量表达转基因拟南芥的根长显著长于WT。【结论】超量表达的MiZAT10A与MiZAT10B可使转基因拟南芥提前开花并提高其对盐、干旱、重金属及外源激素GA₃和ABA的抗性。

关键词: 芒果, 非生物胁迫, 锌指蛋白, 表达模式, 基因功能分析

Abstract:

【Objective】Zinc finger protein (ZFP) plays an important role in plant abiotic stress response. Therefore, to provide a theoretical basis for stress resistance breeding, this study aimed to analyze the response of two zinc finger protein genes of MiZAT10A and MiZAT10B transgenic Arabidopsis to abiotic stresses, such as salt, drought, heavy metals and exogenous hormones. 【Method】 The promoter cis acting elements and motif of mango MiZAT10A and MiZAT10B genes were predicted and analyzed by online software PLACE and MEME, respectively. The chromosome location map was drawn by TBtools software and SiJiMi gene annotation file (GFF file and unpublished). Tissue expression patterns of MiZAT10A and MiZAT10B genes were analyzed by qRT-PCR. The overexpression vectors of MiZAT10A and MiZAT10B genes were constructed and transformed into Arabidopsis thaliana by Agrobacterium floral-dip method. The phenotype of MiZAT10A and MiZAT10B transgenic plant were observed and recorded under salt, drought, heavy metals, abscisic acid and gibberellin treatments. 【Result】 Promoter cis element analysis showed that there were many light response elements, hormone response elements and abiotic stress response elements in the promoter region of MiZAT10A and MiZAT10B genes. Expression analysis showed that MiZAT10A and MiZAT10B were highly expressed in buds and flowers. 9 of MiZAT10A and 14 of MiZAT10B transgenic Arabidopsis strains were obtained. Overexpression of MiZAT10A and MiZAT10B significantly resulted early flowering compared with the control lines. The root length of MiZAT10A and MiZAT10B overexpressing transgenic Arabidopsis was significantly longer than that of control lines under salt stress, drought stress, heavy metal stress, GA₃ and ABA hormone treatments. 【Conclusion】 Overexpression of MiZAT10A and MiZAT10B not only promoted transgenic Arabidopsis flowering early but also improved salt, drought, heavy metals and exogenous hormones GA₃ and ABA resistance.

Key words: mango, abiotic stress, zinc finger protein, expression, function analysis

莫文静,朱嘉伟,何新华,余海霞,江海玲,覃柳菲,张艺粒,李雨泽,罗聪. 芒果MiZAT10A和MiZAT10B功能分析[J]. 中国农业科学, 2023, 56(1): 193-202.

MO WenJing,ZHU JiaWei,HE XinHua,YU HaiXia,JIANG HaiLing,QIN LiuFei,ZHANG YiLi,LI YuZe,LUO Cong. Functional Analysis of MiZAT10A and MiZAT10B Genes in Mango[J]. Scientia Agricultura Sinica, 2023, 56(1): 193-202.

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链接本文: https://www.chinaagrisci.com/CN/10.3864/j.issn.0578-1752.2023.01.015

https://www.chinaagrisci.com/CN/Y2023/V56/I1/193

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引物 Primer	序列 Sequences (5′-3′)	用途 Usage
AtActin2-F	GCAGAGCGGGAAATTGTAAG	半定量Semi-quantitative
AtActin2-R	GTACAGATCCTTCCTGATATCC	半定量Semi-quantitative
MiActin-F	CCGAGACATGAAGGAGAAGC	实时荧光定量qRT-PCR
MiActin-R	GTGGTCTCATGGATACCAGCA	实时荧光定量qRT-PCR
MiZAT10A-F	GCTAAGCGCAAAAGGTCGAA	实时荧光定量qRT-PCR
MiZAT10A-R	GGTCGTAGCAGCTGATGGAG	实时荧光定量qRT-PCR
MiZAT10B-F	GCGТССТСАCAACCATCCAA	实时荧光定量qRT-PCR
MiZAT10B-R	GGAGGTTGACTGCTCGTCG	实时荧光定量qRT-PCR