黄钰莹, 陈颖, 王宝纬, 关范圆, 郑语嫣, 范菁, 王金铃, 胡秀华, 王晓晖. 丹参SmIAA7基因的克隆、亚细胞定位与表达分析J. 药学学报, 2025, 60(2): 514-525. DOI: 10.16438/j.0513-4870.2024-0789
引用本文: 黄钰莹, 陈颖, 王宝纬, 关范圆, 郑语嫣, 范菁, 王金铃, 胡秀华, 王晓晖. 丹参SmIAA7基因的克隆、亚细胞定位与表达分析J. 药学学报, 2025, 60(2): 514-525. DOI: 10.16438/j.0513-4870.2024-0789
HUANG Yu-ying, CHEN Ying, WANG Bao-wei, GUAN Fan-yuan, ZHENG Yu-yan, FAN Jing, WANG Jin-ling, HU Xiu-hua, WANG Xiao-hui. Cloning, subcellular localization and expression analysis of SmIAA7 gene from Salvia miltiorrhizaJ. Acta Pharmaceutica Sinica, 2025, 60(2): 514-525. DOI: 10.16438/j.0513-4870.2024-0789
Citation: HUANG Yu-ying, CHEN Ying, WANG Bao-wei, GUAN Fan-yuan, ZHENG Yu-yan, FAN Jing, WANG Jin-ling, HU Xiu-hua, WANG Xiao-hui. Cloning, subcellular localization and expression analysis of SmIAA7 gene from Salvia miltiorrhizaJ. Acta Pharmaceutica Sinica, 2025, 60(2): 514-525. DOI: 10.16438/j.0513-4870.2024-0789

丹参SmIAA7基因的克隆、亚细胞定位与表达分析

Cloning, subcellular localization and expression analysis of SmIAA7 gene from Salvia miltiorrhiza

  • 摘要: Auxin/indole-3-acetic acid (Aux/IAA) 是植物中一个重要的基因家族, 在生长素信号传导中扮演着关键的角色, 参与了植物的生长、发育以及对逆境胁迫的响应和调节过程。本研究通过分析丹参(Salvia miltiorrhiza) 不同羽状复叶的转录组数据筛选了潜在的与叶片发育相关的基因SmIAA7, 设计特异引物成功从丹参中克隆了SmIAA7, 并进行生物信息学分析、原核表达和纯化、亚细胞定位、生长素及非生物胁迫下的诱导表达分析。SmIAA7基因全长684 bp, 编码227个氨基酸, 蛋白分子质量25.3 kD。保守结构域分析表明SmIAA7含有超家族保守结构域Aux_IAA (pfam02309), 序列分析及系统进化树结果表明SmIAA7和其他植物中的IAA7以及IAA14同源性较高, 可能参与植物的生长发育并响应逆境胁迫。构建原核表达载体pET28a-SmIAA7并在大肠杆菌Rosetta (DE3) 菌株中成功表达SmIAA7重组蛋白, 纯化得到可溶性重组蛋白。亚细胞定位实验结果表明SmIAA7定位于植物细胞的细胞核中。实时荧光定量PCR结果表明, SmIAA7基因能够响应生长素的诱导导致表达水平升高。干旱、低温以及盐胁迫均能够显著提高SmIAA7基因的表达水平。本研究为进一步揭示SmIAA7在丹参叶片发育、信号传递及逆境防御中的作用奠定基础。

     

    Abstract: The auxin/indole-3-acetic acid (Aux/IAA) gene family is an important regulator for plant growth hormone signaling, involved in plant growth, development, as well as response to environmental stresses. In the present study, we identified SmIAA7 which is potentially associated with Salvia miltiorrhiza leaf development through comparatively analyzed the transcriptome data from different pinnate leaves. SmIAA7 was successfully isolated from S. miltiorrhiza using the specific primers. Then subsequent bioinformatic analysis, prokaryotic expression and purification, subcellular localization, and induction expression analysis under auxin and abiotic stress were performed. The full-length of SmIAA7 contained an ORF of 684 bp encoding a protein of 227 amino acid with a molecular weight of 25.3 kD. Conserved domain analysis showed that SmIAA7 contains the conserved Aux_IAA domain (pfam02309). Sequence analysis and phylogenetic tree analysis results indicated SmIAA7 was phylogenetically close to IAA7 and IAA14 from other plants, suggesting SmIAA7 involved in plant growth and development as well as response to environmental stresses. The prokaryotic expression vector pET28a-SmIAA7 was constructed and SmIAA7 recombinant protein was successfully expressed in E. coli Rosetta (DE3) strain. Subcellular localization experiment demonstrated that SmIAA7 localized in the nucleus of plant cells. Real-time fluorescence quantitative PCR results showed that the expression level of SmIAA7 was upregulated in response to auxin. Drought, low temperature, and salt stress significantly increased the transcript level of SmIAA7 gene. This study lays a foundation for further elucidating the role of SmIAA7 in leaf development, signal transduction, and stress defense in S. miltiorrhiza.

     

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