扁蓿豆<i>MrAGL8</i>基因的克隆、亚细胞定位及表达分析

doi:10.7668/hbnxb.20195685

华北农学报 ›› 2025, Vol. 40 ›› Issue (5): 55-61. doi: 10.7668/hbnxb.20195685

所属专题： 生物技术；

• 作物遗传育种·种质资源·生物技术 • 上一篇下一篇

扁蓿豆MrAGL8基因的克隆、亚细胞定位及表达分析

于佳¹^,²^,³, 郭慧琴¹^,²^,³^,, 李宇霞¹^,²^,³, 雷慧¹^,²^,³, 任卫波⁴

¹ 内蒙古农业大学生命科学学院植物生物技术功能研究室, 内蒙古呼和浩特 010011
² 麦类种质创新利用自治区高等学校重点实验室, 内蒙古呼和浩特 010011
³ 内蒙古自治区旱寒区植物逆境适应与遗传修饰改良重点实验室, 内蒙古呼和浩特 010011
⁴ 内蒙古大学生态与环境学院, 内蒙古呼和浩特 010020

收稿日期:2024-12-16 出版日期:2025-11-03
通讯作者:
郭慧琴(1979—),女,内蒙古巴彦淖尔人,副教授,硕士,硕士生导师,主要从事植物分子生物学研究。
作者简介:
于佳(2000—),女,黑龙江哈尔滨人,在读硕士,主要从事植物分子生物学研究。
基金资助:
内蒙古自治区种业科技创新重大示范工程“揭榜挂帅”项目(2022JBGS0040); “科技兴蒙”重点专项(2020-科技兴蒙-草种业技术创新中心-2)

Cloning,Subcellular Localization and Expression Analysis of MrAGL8 Gene in Medicago ruthenica

YU Jia¹^,²^,³, GUO Huiqin¹^,²^,³^,, LI Yuxia¹^,²^,³, LEI Hui¹^,²^,³, REN Weibo⁴

¹ Plant Biotechnology Function Laboratory, College of Life Sciences,Inner Mongolia Agricultural University, Hohhot 010011 China
² Key Laboratory of Wheat Germplasms Innovation and Utilization Autonomous Region Higher School, Hohhot 010011 China
³ Key Laboratory of Plant Stress Adaptation and Genetic Modification Improvement in Arid and Cold Regions of Inner Mongolia Autonomous Region, Hohhot 010011 China
⁴ School of Ecology and Environment, Inner Mongolia University, Hohhot 010020 China

Received:2024-12-16 Published:2025-11-03

摘要/Abstract

摘要： 为了探究扁蓿豆MrAGL8基因与裂荚性状间的联系。以扁蓿豆为植物材料,利用PCR技术扩增MrAGL8基因并克隆测序,并对该基因进行生物信息学分析、亚细胞定位和不同组织器官的表达分析。结果表明,利用PCR扩增技术克隆获得了长度为711 bp的MrAGL8 cDNA完整编码区。生物信息学分析显示,MrAGL8基因编码236个氨基酸,MrAGL8包含MADS-box和K-box蛋白结构域,预测其蛋白质分子质量为27.39 ku,理论等电点为8.69,带正电氨基酸残基总数为39个,带负电氨基酸残基总数为36个,不稳定系数为48.5。预测蛋白质二级结构包含α-螺旋和β-转角,属于不稳定的亲水碱性蛋白。亚细胞定位分析显示,MrAGL8定位于细胞核中。不同组织器官的表达分析结果显示,MrAGL8在不同组织中的表达量为茎>根>荚果>叶>花,且根和茎与叶、花、荚果的表达量差异显著。结果表明,扁蓿豆MrAGL8基因与荚果开裂相关。

关键词: 扁蓿豆, MrAGL8, 基因克隆, 亚细胞定位, 表达分析

Abstract:

In order to explore the relationship between MrAGL8 gene and pod dehiscence traits in Medicago ruthenica. In this study,Medicago ruthenica was used as the plant material.The MrAGL8 gene was amplified by PCR,cloned,and sequenced.Additionally,bioinformatics analysis,subcellular localization,and expression analysis in different tissues and organs were performed for this gene.The results showed that the complete coding region of MrAGL8 cDNA with a length of 711 bp was obtained through cloning using PCR amplification technology.Bioinformatics analysis results showed that MrAGL8 encoded 236 amino acids,it had MADS-box and K-box protein conserve domains.Its molecular weight was 27.39 ku,the theoretical isoelectric point was 8.69,the total number of positively charged residues was 39,the total number of negatively charged residues was 36,and the instability coefficient was 48.5.The secondary structure of the protein contained α-helix and β-sheet.It was an unstable protein and belonged to a hydrophilic alkaline protein.Subcellular localization results showed that MrAGL8 protein was located in the nucleus.The expression analysis of different tissues and organs showed that the expression level of MrAGL8 in different tissues was stem>root>pod>leaf>flower,and the expression levels of roots and stems were significantly different from those of leaves,flowers and pods.The results showed that MrAGL8 gene was related to pod dehiscence.

Key words: Medicago ruthenica, MrAGL8, Gene cloning, Subcellular localization, Expression analysis

中图分类号:

S541.9

于佳, 郭慧琴, 李宇霞, 雷慧, 任卫波. 扁蓿豆MrAGL8基因的克隆、亚细胞定位及表达分析[J]. 华北农学报, 2025, 40(5): 55-61. doi: 10.7668/hbnxb.20195685.

YU Jia, GUO Huiqin, LI Yuxia, LEI Hui, REN Weibo. Cloning,Subcellular Localization and Expression Analysis of MrAGL8 Gene in Medicago ruthenica[J]. Acta Agriculturae Boreali-Sinica, 2025, 40(5): 55-61. doi: 10.7668/hbnxb.20195685.

http://www.hbnxb.net/CN/Y2025/V40/I5/55

图/表 10

图1 MrAGL8 CDS的PCR扩增电泳检测

Fig.1 PCR amplification and electrophoresis detection of MrAGL8 CDS M.Trans 2K DNA Ladder;1—4.PCR产物。 M.Trans 2K DNA Ladder;1—4.PCR products.

图2 MrAGL8 CDS克隆菌液PCR扩增和电泳检测

Fig.2 PCR amplification and electrophoresis detection of MrAGL8 CDS colonies M.Trans 2K DNA Ladder;1—8.PCR 产物。 M.Trans 2K DNA Ladder;1—8.PCR products.

图3 MrAGL8 cDNA编码区序列及编码氨基酸序列

Fig.3 MrAGL8 cDNA coding region sequence and coded amino acid sequence

图4 MrAGL8 结构域预测

Fig.4 Domain prediction of MrAGL8

图5 MrAGL8的系统发育树

Fig.5 Phylogenetic tree of MrAGL8

图6 MrAGL8 亲/疏水性分析

Fig.6 Hydrophilic or hydrophobic analysis of MrAGL8

图7 MrAGL8蛋白质二级结构预测蓝色.α-螺旋;绿色.β-转角;紫色.延伸链;橙色.无规则卷曲。

Fig.7 Secondary structure prediction of MrAGL8 protein Blue.α-helix;Green.β-turn;Purple.Extended strand;Orange.Random coil.

图8 MrAGL8蛋白三级结构预测

Fig.8 Tertiary structure prediction of MrAGL8 protein

图9 MrAGL8在烟草叶片下表皮细胞中亚细胞定位 Bright.明场;GFP.绿色荧光通道;DAPI(4,6-二氨基-2-苯基吲哚).细胞核定位阳性对照;Merged.叠加场。

Fig.9 Subcellular localization of MrAGL8 in lower epidermal cells of tobacco leaves Bright.Bright field;GFP.Green fluorescence channel;DAPI(4,6-diamidino-2-phenylindole). Positive control for nuclear localization;Merged.Mixed field

图10 MrAGL8基因在不同组织中的相对表达量不同小写字母表示不同处理间差异显著(P<0.05)。

Fig.10 The relative expression of MrAGL8 gene in different tissues Different lowercase letters indicate significant differences in different treatments(P<0.05).

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扁蓿豆MrAGL8基因的克隆、亚细胞定位及表达分析

Cloning,Subcellular Localization and Expression Analysis of MrAGL8 Gene in Medicago ruthenica

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