大豆抗胞囊线虫锌指蛋白基因GmC_(2)H_(2)-2like克隆与初步功能分析  

Soybean Cyst Nematode Resistant Zinc Finger Protein Gene GmC_(2)H_(2)-2like Cloning and Preliminary Functional Analysis

作　　者：曲硕 刘芳 宋庚晨 胡世豪 房瑶瑶 赵雪[1] 韩英鹏[1] QU Shuo;LIU Fang;SONG Gengchen;HU Shihao;FANG Yaoyao;ZHAO Xue;HAN Yingpeng(College of Agriculture,Northeast Agricultural University,Northeast Key Laboratory of Soybean Biology and Genetic Breeding,Ministry of Agriculture and Rural Affairs,Harbin 150030,China)

机构地区：[1]东北农业大学农学院,农业农村部东北大豆生物学与遗传育种重点实验室,黑龙江哈尔滨150030

出　　处：《华北农学报》2024年第3期195-207,共13页Acta Agriculturae Boreali-Sinica

基　　金：黑龙江省重点基金项目(ZD2022C002);黑龙江省重点研发项目(JD22A015);国家自然科学基金项目(U22A20473);国家现代农业岗位体系项目(CARS-04-PS07);东北农业大学科研项目(NEAU2023QNLJ-003)。

摘　　要：为了研究锌指蛋白C_(2)H_(2)在大豆胞囊线虫病(SCN)胁迫应答中起到的重要作用,为进一步研究抗大豆胞囊线虫奠定重要基础。利用东农L-10(抗SCN)、黑农37(感SCN)接种SCN3根系转录组数据,筛选出差异表达基因GmC_(2)H_(2)-2like。倒置荧光显微镜下观察该GmC_(2)H_(2)-2like定位情况,对该基因进行生物信息学分析,克隆并构建其超表达载体转化大豆毛状根,研究其对胞囊线虫病的抗性功能。结果表明,GmC_(2)H_(2)-2like编码410个氨基酸残基,分子质量为45.77 ku,等电点为8.73,含47个磷酸化位点,蛋白二级结构含α-螺旋(25.61%)、无规则卷曲(57.32%)、延伸链(13.66%)和β-转角(3.14%)4种结构;亚细胞定位结果发现,其编码蛋白位于细胞核;分析超表达转GmC_(2)H_(2)-2like大豆毛状根发现,其根部组织的单位面积胞囊线虫数量显著低于空载体对照,表明GmC_(2)H_(2)-2like具有抑制胞囊线虫的功能。In order to study the important role of zinc finger protein C_(2)H_(2)in the stress response to soybean cyst nematode disease(SCN),lay an important foundation for further research on anti-soybean cyst nematode.The transcriptome data of SCN3 roots inoculated with Dongnong L-10(SCN-resistant)and Heinong 37(SCN-susceptible)were utilized in the present study,and the differentially expressed gene,GmC_(2)H_(2)-2like,was screened out.The localization of this gene was observed under the inverted fluorescence microscope.The gene was bioinformatically analyzed,and its overexpression vector was cloned and constructed to transform soybean hairy roots to study its resistance function against cyst nematode disease.The results showed that GmC_(2)H_(2)-2like encoded 410 amino acid residues,with a molecular weight of 45.77 ku,an isoelectric point of 8.73,and 47 phosphorylation sites,and the protein tertiary structure contained four structures,namely,α-helix(25.61%),random coil(57.32%),extended strand(13.66%),andβ-turn(3.14%);the results of the subcellular localization revealed that the encoded protein was located in the nucleus;analysis of the overexpressed trans-GmC_(2)H_(2)-2like soybean hairy roots revealed that the number of cyst nematodes per unit area of the root tissue was significantly lower than that of the empty vector control,suggesting that GmC_(2)H_(2)-2like has the function of inhibiting cyst nematodes.

关 键 词：大豆 胞囊线虫 GmC_(2)H_(2)-2like 生物信息学 亚细胞定位 

分 类 号：S435.651[农业科学—农业昆虫与害虫防治]