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作物学报 ›› 2026, Vol. 52 ›› Issue (4): 1116-1126.doi: 10.3724/SP.J.1006.2026.54114

• 作物遗传育种·种质资源·分子遗传学 • 上一篇    下一篇

马铃薯Argonaute基因家族鉴定及晚疫病诱导表达分析

田立涛1,2(), 丁宁2,3, 王树林2,3, 齐恩芳2,3, 张荣2,3, 王蕊蕊2, 马利雯2, 李建武1,2,3,*(), 杨江伟1,*()   

  1. 1甘肃农业大学生命科学技术学院, 甘肃兰州 730070
    2甘肃省农业科学院马铃薯研究所 / 甘肃省马铃薯种质资源创新工程实验室, 甘肃兰州 730070
    3国家种质资源渭源观测实验站, 甘肃渭源 748201
  • 收稿日期:2025-09-25 接受日期:2026-01-22 出版日期:2026-04-12 网络出版日期:2026-02-05
  • 通讯作者: *李建武, E-mail: ljw0931@163.com; 杨江伟, E-mail: yjw@gsau.edu.cn
  • 作者简介:E-mail: TianLT12@outlook.com
  • 基金资助:
    甘肃省农业科学院现代生物育种专项(2025GAAS15);甘肃省农业农村厅种业公关项目(ZYGG-2025-4);国家现代农业产业技术体系(CARS-09-P06);国家自然科学基金项目(31760410)

Genome-wide identification of the Argonaute gene family and its induction by late blight in potato (Solanum tuberosum L.)

Tian Li-Tao1,2(), Ding Ning2,3, Wang Shu-Lin2,3, Qi En-Fang2,3, Zhang Rong2,3, Wang Rui-Rui2, Ma Li-Wen2, Li Jian-Wu1,2,3,*(), Yang Jiang-Wei1,*()   

  1. 1College of Life Science and Technology, Gansu Agricultural University, Lanzhou 730070, Gansu, China
    2Potato Research Institute of Gansu Academy of Agricultural Sciences / Gansu Engineering Laboratory of Potato Germplasm Resources Innovation, Lanzhou 730070, Gansu, China
    3National Germplasm Resources Agricultural Experimental Station, Weiyuan 748201, Gansu, China
  • Received:2025-09-25 Accepted:2026-01-22 Published:2026-04-12 Published online:2026-02-05
  • Contact: *E-mail: ljw0931@163.com; E-mail: yjw@gsau.edu.cn
  • Supported by:
    Modern Biological Breeding Program of Gansu Academy of Agricultural Sciences(2025GAAS15);Seed Industry Research Project of Gansu Provincial Department of Agriculture and Rural Affairs(ZYGG-2025-4);China Agriculture Research System(CARS-09-P06);National Natural Science Foundation of China(31760410)

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摘要：

Argonaute (AGO)蛋白作为RNA诱导沉默复合体(RNA-induced silencing complex, RISC)的核心元件, 在小RNA (sRNA)介导的基因沉默中具有重要作用。在植物中, AGO蛋白广泛参与调控发育、胁迫应答及病原防御等关键生物学过程, 然而目前在马铃薯中对该基因家族的系统研究尚无报道。本研究通过生物信息学方法对StAGO基因家族成员进行鉴定, 并分析了不同组织和致病疫霉菌(Phytophthora infestans (Mont.) de Bary)侵染后叶片的表达, 同时利用拟南芥原生质体瞬时表达体系验证了StAGO1A和StAGO2A在病原胁迫下的应答。结果表明, 在双单倍体马铃薯全基因组中共筛选出14个StAGO基因, 分布在7条染色体上。根据系统进化关系可将其分为3个亚家族: StAGO1A/ StAGO1B/StAGO5/StAGO10/StAGO10A、StAGO2A/StAGO2B/StAGO3/StAGO7和StAGO4A/StAGO4B/StAGO4/StAGO6/ StAGO8。共线性分析结果显示, 该基因家族成员间存在2个片段复制事件(StAGO1A和StAGO1B, StAGO4A和StAGO4B)。通过RT-qPCR分析StAGOs基因在不同组织中的表达谱发现, StAGO10在叶中高表达, StAGO1B在茎中高表达, 而StAGO3、StAGO5、StAGO7等在试管薯中相对表达量高。在晚疫病病原菌侵染离体叶片后, StAGO1A、StAGO2A、StAGO3、StAGO5、StAGO6、StAGO10均显著上调表达。此外, 本研究构建了瞬时表达载体在拟南芥原生质体中进行体内表达验证, 发现晚疫病病原菌能诱导StAGO1A与StAGO2A表达增强。综上所述, 本研究为进一步分析StAGOs基因家族功能及其在响应马铃薯晚疫病中的作用奠定了理论基础。

关键词: 马铃薯, Argonaute基因, 全基因组鉴定, 表达分析, 晚疫病

Abstract:

Argonaute (AGO) proteins are core components of the RNA-induced silencing complex (RISC) and play a crucial role in small RNA (sRNA)-mediated gene silencing. In plants, AGO proteins regulate key biological processes such as development, stress responses, genome architecture, and pathogen defense. However, the AGO gene family in potato (Solanum tuberosum L.) has not yet been characterized. In this study, we identified members of the StAGO gene family through bioinformatics approaches and analyzed their expression patterns in various tissues and in leaves infected by Phytophthora infestans (Mont.) de Bary using real-time quantitative PCR (RT-qPCR). The expression of StAGO1A and StAGO2A under pathogen stress was further validated using a transient expression assay in Arabidopsis protoplasts. A total of 14 StAGO genes were identified in the doubled haploid potato genome, distributed across seven chromosomes. Phylogenetic analysis grouped them into three subfamilies: StAGO1A, StAGO1B, StAGO5, StAGO10, StAGO10A; StAGO2A, StAGO2B, StAGO3, StAGO7; and StAGO4A, StAGO4B, StAGO4, StAGO6, StAGO8. Collinearity analysis revealed two segmental duplication events within the family (StAGO1A and StAGO1B, StAGO4A and StAGO4B). Tissue-specific expression analysis by RT-qPCR showed that StAGO10 was highly expressed in leaves, StAGO1B showed elevated expression in stems, and StAGO3, StAGO5, and StAGO7 exhibited relatively high expression in mini tubers. Moreover, expression profiling following infection with the late blight pathogen revealed significant upregulation of StAGO1A, StAGO2A, StAGO3, StAGO5, StAGO6, and StAGO10. Transient expression assays in Arabidopsis protoplasts further confirmed that P. infestans infection induces the expression of StAGO1A and StAGO2A. In conclusion, this study provides a theoretical foundation for future functional analyses of the StAGO gene family and their roles in potato defense against late blight.

Key words: potato, Argonaute (AGO) gene, genome-wide identification, expression analysis, late blight

图1

‘大西洋’组培苗"

表1

本研究所用引物"

引物名称
Primer name		 引物序列
Primer sequence (5'-3')		 说明
Describe
StEF1α-F ATTGGAAACGGATATGCTCCA 马铃薯内参引物
StEF1α-R TCCTTACCTGAACGCCTGTCA Potato reference gene primers
StAGO1A-F ACCACCTTGGCTTTGACCTT StAGO1A RT-qPCR引物
StAGO1A-R AATGGCGCATAATCACCCGA StAGO1A RT-qPCR primers
StAGO1B-F ATCTGAGCCCCCTGAGATGT StAGO1B RT-qPCR引物
StAGO1B-R TGTCAGGTAGCTCCGCAAAG StAGO1B RT-qPCR primers
StAGO10-F GAACCACTCCCGGTTGTTGA StAGO10 RT-qPCR引物
StAGO10-R TAGCTCCCTTGTTGGCTGTG StAGO10 RT-qPCR primers
StAGO10A-F AAGATTGGCCTGAGGTGACG StAGO10A RT-qPCR引物
StAGO10A-R CTTTTGGCCTGTGGCCTTTC StAGO10A RT-qPCR primers
StAGO5-F CCCAATGTTGCCACCCAAAG StAGO5 RT-qPCR引物
StAGO5-R AGAATCTTCTCCCGGTTGCG StAGO5 RT-qPCR primers
StAGO7-F TCGCATGGCATGTTCGAGAT StAGO7 RT-qPCR引物
StAGO7-R TTACACCCCAATGGCTGCAT StAGO7 RT-qPCR primers
StAGO2A-F CCGTCGGACATAGCAAAGGT StAGO2A RT-qPCR引物
StAGO2A-R TGAAAGAGAGACATCAAATACCTCA StAGO2A RT-qPCR primers
StAGO2B-F TGGTTGACGGATAAGCCTGC StAGO2B RT-qPCR引物
StAGO2B-R GGACAACGCCATCCCTTGTA StAGO2B RT-qPCR primers
StAGO3-F GTCTCCGGGTACTGTTGTGG StAGO3 RT-qPCR引物
StAGO3-R GACGACGAAGCTTCAGAGGA StAGO3 RT-qPCR primers
StAGO4A-F TGCCTCTTTTGAGCAGAAGAATTT StAGO4A RT-qPCR引物
StAGO4A-R CCCTTGTTTCCAACACCACG StAGO4A RT-qPCR primers
StAGO4B-F GCTTAGAAGGGCTCCACCTG StAGO4B RT-qPCR引物
StAGO4B-R AACTCTTCCAGGAGCCAAGC StAGO4B RT-qPCR primers
StAGO4-F GAAATCCTGCTCCTCTGGGC StAGO4 RT-qPCR引物
StAGO4-R AGCTGCTCGATAGCGTGAAA StAGO4 RT-qPCR primers
StAGO8-F TGGGACTAGCAGGCCAATTC StAGO8 RT-qPCR引物
StAGO8-R TGCCGATTTCTTAGGGCCTTT StAGO8 RT-qPCR primers
StAGO6-F ACTGTTTTAGGGCTGGGCAA StAGO6 RT-qPCR引物
StAGO6-R GCCCTCACCAAGATGGTTGT StAGO6 RT-qPCR primers
AGO2A-PstI-F ACTCTAGAAAGCTTCTGCAGATGGACCGTGGGAACTACCG StAGO2A CDS序列扩增引物
AGO2A-SpeI-R TACCGGATCCACTAGTGACGAAAAACATTATGTCCTGC StAGO2A CDS sequence clone primers
AGO1A-PstI-F ACTCTAGAAAGCTTCTGCAGATGGTGCGGAAGAGGAGAAC StAGO1A CDS序列扩增引物
AGO1A-SpeI-R TACCGGATCCACTAGTACAATAGAACATCACCCTTTTG StAGO1A CDS sequence clone primers

表2

StAGOs蛋白的理化性质"

名称
Name		 基因ID
Gene ID		 氨基酸
Amino acid (aa)		 分子量
Molecular weight (kD)		 等电点
pI		 不稳定系数
Instability index		 亲水性均值
GRAVY
StAGO1A Soltu.DM.06G027550 1054 117.03 9.46 50.28 -0.519
StAGO1B Soltu.DM.03G019130 1127 124.70 9.48 49.97 -0.598
StAGO10 Soltu.DM.12G024630 945 106.95 9.25 48.24 -0.424
StAGO10A Soltu.DM.09G025190 982 110.56 9.30 44.96 -0.444
StAGO5 Soltu.DM.06G030090 1025 114.25 9.39 49.08 -0.484
StAGO7 Soltu.DM.01G010020 1002 114.24 9.20 47.28 -0.411
StAGO2A Soltu.DM.02G012280 1051 117.08 9.30 41.75 -0.560
StAGO2B Soltu.DM.02G012290 821 92.68 7.98 34.58 -0.396
StAGO3 Soltu.DM.02G012300 999 112.34 9.18 39.95 -0.571
StAGO4A Soltu.DM.01G005850 909 101.75 9.01 47.80 -0.399
StAGO4B Soltu.DM.06G028860 913 101.94 8.95 51.29 -0.405
StAGO4 Soltu.DM.01G035930 809 90.51 9.23 41.79 -0.431
StAGO8 Soltu.DM.03G025140 907 102.56 9.49 41.78 -0.317
StAGO6 Soltu.DM.07G016700 953 107.21 8.58 42.53 -0.322

图2

马铃薯AGOs蛋白的进化树和AGOs基因的共线性分析 A: 进化树; 粉色圆点: StAGO基因家族; 蓝色圆点: AtAGO基因家族; 绿色圆点: SlAGO基因家族; Cluster 1: 亚家族I; Cluster 2: 亚家族II; Cluster 3: 亚家族III。B: 共线性分析; St1-St12: 马铃薯的12条染色体; At1-At5: 拟南芥的5条染色体; 灰线: 马铃薯与拟南芥物种间共线性关系; 红线: 目的基因与拟南芥物种间共线性关系; 绿线: 马铃薯物种内候选基因的共线性关系。"

图3

马铃薯AGOs基因家族成员的基因结构和蛋白结构域 A: 基因结构。B: 蛋白结构域。"

图4

StAGOs蛋白PIWI结构域多序列比对 红色: DDH/D催化三联体; 蓝色: 第798位组氨酸(H798); 绿色: 保守QF-V基序。"

图5

StAGOs基因组织表达模式分析 RT-qPCR数据为3次生物学重复、3次技术重复的平均值(n = 9); 不同小写字母代表处理组间差异显著(P < 0.05, Tukey多重比较)。"

图6

陇薯19号(A)和CIP309056.5 (B)侵染晚疫病后StAGOs基因的相对表达量 Mock: 对照组; P. infestans: 晚疫病侵染处理组。RT-qPCR数据为3次生物学重复、3次技术重复的平均值(n = 9)。*和**分别表示在0.05和0.01水平差异显著。"

图7

拟南芥原生质体瞬时表达 A: 拟南芥原生质体瞬时表达; pSuper1300-eGFP: 空载体; pSuper1300-StAGO1A-eGFP: AGO1A过表达载体; pSuper1300-StAGO2A- eGFP: AGO2A过表达载体; Mock: 对照组; P. infestans: 晚疫病侵染处理组; 比例尺: 50 μm。B: 绿色荧光信号统计图; **表示在0.01水平差异显著; 代表性图片来自3次生物学重复(n = 3)。"

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