甜樱桃<i>PavJAZ1</i>基因鉴定、克隆及在果实中的表达分析

doi:10.7668/hbnxb.20195818

摘要/Abstract

摘要：

茉莉酸类物质在调控果实成熟和品质形成过程中发挥着重要作用。JAZ蛋白是茉莉酸信号转导通路的核心抑制因子。为研究茉莉酸信号转导关键抑制因子JAZ蛋白在甜樱桃果实成熟和品质调控中的功能,从甜樱桃基因组中鉴定并克隆了PavJAZ1基因,基于生物信息学,分析了其理化性质、蛋白结构、保守结构域分布、启动子顺式作用元件等信息,通过荧光定量PCR分析了其在甜樱桃果实中的表达模式,观察了PavJAZ1蛋白亚细胞定位;此外,借助双分子荧光互补(BiFC)技术验证了PavJAZ1与PavMYC2之间的互作关系。基因鉴定与克隆结果表明,PavJAZ1基因开放阅读框长度为837 bp,编码278个氨基酸,蛋白分子质量为30.09 ku,理论等电点为9.21,不稳定系数为54.14,亲水性系数为-0.564,说明其为碱性亲水蛋白,且蛋白性质不稳定。蛋白序列和结构分析表明,PavJAZ1主要由无规则卷曲和α-螺旋构成,包含Tify和Jas 2个保守结构域,且与其他物种同源性较高,其中与同为蔷薇科李属的桃同源性最高。亚细胞定位观察发现,PavJAZ1蛋白定位于细胞核。表达模式分析表明,随着果实的发育和成熟,PavJAZ1表达量呈现先升高后逐步下降的趋势,暗示其可能在果实不同阶段发挥不同的作用。此外,BiFC结果表明,PavJAZ1可以与PavMYC2在植物体内互作,初步证明其可以参与茉莉酸信号转导。

关键词: 甜樱桃, JAZ, 果实成熟, 表达模式, 果实品质

Abstract:

Jasmonates play a crucial role in regulating fruit ripening and quality formation.The JAZ proteins are key repressors of the jasmonic acid signaling pathway.To investigate the function of PavJAZ1 in sweet cherry fruit ripening and quality regulation,the PavJAZ1 gene was identified and cloned from the sweet cherry genome.Bioinformatics analysis was performed to explore its physicochemical properties,protein structure,conserved domains,and cis-acting elements in the promoter region.The gene expression pattern of PavJAZ1 in fruits was analyzed by Quantitative Real-time PCR.The subcellular localization of the PavJAZ1 protein was observed,and the interaction between PavJAZ1 and PavMYC2 was verified using the bimolecular fluorescence complementation(BiFC).The results of gene identification and cloning showed that the open reading frame of the PavJAZ1 gene was 837 bp in length,encoding 278 amino acids.The molecular weight of the PavJAZ1 was 30.09 ku,the theoretical isoelectric point was 9.21,the instability index was 54.14,and the grand average of hydropathicity was -0.564,indicating that it was an alkaline hydrophilic and unstable properties.Protein structure analysis revealed that PavJAZ1 was mainly composed of random coils and α-helices,containing two conserved domains,the Tify domain and the Jas domain.It also exhibited high homology with proteins from other species,among which it showed the highest homology with that of peach,a plant that also belonged to the genus Prunus of the Rosaceae family.Subcellular localization analysis showed that the PavJAZ1 protein was localized in the nucleus.The expression level of PavJAZ1 increased significantly in the early stages of fruit development and then gradually decreased.This result suggested that PavJAZ1 may play different roles at different developmental stages.In addition,the BiFC results showed that PavJAZ1 interacted with PavMYC2,confirming that PavJAZ1 participates in the jasmonic acid signaling pathway.

Key words: Sweet cherry, JAZ, Fruit ripening, Expression pattern, Fruit quality

中图分类号:

王未, 冉江林, 张晓明, 闫国华, 王晶, 周宇, 吴传宝, 冯琛, 张开春, 段续伟. 甜樱桃PavJAZ1基因鉴定、克隆及在果实中的表达分析[J]. 华北农学报, 2026, 41(2): 28-35. doi: 10.7668/hbnxb.20195818.

WANG Wei, RAN Jianglin, ZHANG Xiaoming, YAN Guohua, WANG Jing, ZHOU Yu, WU Chuanbao, FENG Chen, ZHANG Kaichun, DUAN Xuwei. Identification,Cloning and Expression Analysis of PavJAZ1 in the Fruit of Prunus avium L.[J]. Acta Agriculturae Boreali-Sinica, 2026, 41(2): 28-35. doi: 10.7668/hbnxb.20195818.

http://www.hbnxb.net/CN/Y2026/V41/I2/28

图/表 9

图1 PavJAZ1基因扩增 M.DNA marker;1,2.目的片段。

Fig.1 PCR amplification of PavJAZ1 gene M.DNA marker;1,2.Target fragment.

图2 PavJAZ1与其他物种JAZ1进化分析

Fig.2 Phylogenetic analysis of PavJAZ1 and JAZ1 in other species

图3 PavJAZ1保守结构域分析及序列比对 A.PavJAZ1保守结构域分析;B.PavJAZ1保守结构域序列比对。

Fig.3 Conserved domain analysis and sequence alignment of PavJAZ1 A.Conserved domain analysis of PavJAZ1; B.Conserved domain sequence alignment of PavJAZ1.

图4 PavJAZ1基因结构分析及编码蛋白结构预测 A. PavJAZ1基因结构分析;B.PavJAZ1蛋白二级结构预测;C.PavJAZ1蛋白三级结构预测。

Fig.4 Structure analysis of PavJAZ1 and prediction of the structure of the encoded protein A.Gene structure analysis of PavJAZ1;B.Secondary structure prediction of PavJAZ1;C.Tertiary structure prediction of PavJAZ1.

表1 PavJAZ1基因启动子主要顺式作用元件

Tab.1 PavJAZ1 gene promoter cis-acting elements

分类 Type	名称 Name	序列 Sequence	个数 Number	功能 Function
胁迫应答元件	DRE core	GCCGAC	1	干旱应答元件
Stress related element	3-AF1 binding site	TAAGAGAGGAA	1	光应答元件
	Box 4	ATTAAT	4	光应答元件
	G-Box	CACGTG	5	光应答元件
	GATA-motif	AAGGATAAGG	1	光应答元件
	Sp1	GGGCGG	2	光应答元件
	TCCC-motif	TCTCCCT	1	光应答元件
	GC-motif	CCCCCG	1	缺氧特异性诱导元件
	TC-rich repeats	ATTCTCTAAC	1	防御和应激调控元件
	WUN-motif	CCATTTCAA	1	创伤响应元件
激素应答元件	ABRE	ACGTG	4	脱落酸应答元件
Hormone response element	TGACG-motif	TGACG	1	茉莉酸应答元件
	CGTCA-motif	CGTCA	1	茉莉酸应答元件
	P-box	CCTTTTG	1	赤霉素应答元件
	TATC-box	TATCCCA	1	赤霉素应答元件
	TCA-element	CCATCTTTTT	1	水杨酸应答元件
	TGA-element	AACGAC	1	生长素应答元件
	ERE	ATTTCATA	2	乙烯应答元件

图5 PavJAZ1蛋白亚细胞定位分析

Fig.5 Subcellular localization of PavJAZ1 protein

图6 PavJAZ1在果实不同发育时期表达分析不同小写字母表示差异显著(P<0.05)。

Fig.6 Expression analysis of PavJAZ1 in different fruit developmental stage Different lowercase letters indicate significant differences(P<0.05).

图7 PavJAZ1互作蛋白预测 A.基于桃互作蛋白预测;B.基于拟南芥互作蛋白预测。

Fig.7 Predicted proteins interacting with PavJAZ1 A.Prediction based on protein-protein interactions in peach;B.Prediction based on protein-protein interactions in Arabidopsis.

图8 PavJAZ1与PavMYC2蛋白互作验证

Fig.8 Analysis of the interaction between PavJAZ1 and PavMYC2

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甜樱桃PavJAZ1基因鉴定、克隆及在果实中的表达分析

Identification,Cloning and Expression Analysis of PavJAZ1 in the Fruit of Prunus avium L.

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甜樱桃PavJAZ1基因鉴定、克隆及在果实中的表达分析

Identification,Cloning and Expression Analysis of PavJAZ1 in the Fruit of Prunus avium L.

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