抗性候选基因GmMIOX4的表达分析、克隆及过表达载体的构建  

Resistance Candidate Genes Gm MIOX4 Gene Expression Analysis,Cloning and Over Expression Vector Constructing

作　　者：刘婷[1] 杨若巍 王学敏[1] 于佰双[2] 王惠[1] 段玉玺[3] LIU Ting YANG Ruo-wei WANG Xue-min YU Bai-shuang WANG Hui DUAN Yu-xi(Shenyang Agricultural University, College of Bioscience and Bioteehnology, Shenyang 110866, China Soybean Research Institute, Heilongjiang Academy of Agricultural Sciences, Harbin 150086, China Shenyang Agricultural University, College of Plant Protection, Shenyang 110866, China)

机构地区：[1]沈阳农业大学生物科学技术学院,辽宁沈阳110866 [2]黑龙江省农业科学院大豆研究所,黑龙江哈尔滨150086 [3]沈阳农业大学植物保护学院,辽宁沈阳110866

出　　处：《大豆科学》2016年第5期730-735,共6页Soybean Science

基　　金：国家自然科学基金重点项目(31330063)

摘　　要：将两个不同抗性大豆品种灰皮支黑豆和辽豆15人工接种大豆胞囊线虫3号生理小种(SCN3),利用实时荧光定量PCR技术,对SCN3侵染后的早期和线虫完成一个整个生活史时的Gm MIOX4基因相对表达量变化进行分析。结果表明:在SCN3侵染下,辽豆15根内的Gm MIOX4基因在25 dpi时,相对表达量升高,而灰皮支黑豆根内的Gm MIOX4基因在10 dpi时被诱导表达,相对表达量达到最高值,表明该基因在此时期发挥一定作用。线虫侵入大豆根内10 d是其通过合胞体吸取寄主营养从而维持正常生长发育的时期,在10 dpi该基因被诱导表达,使其合胞体结构发生改变,从而抑制线虫发育,最终导致死亡或使灰皮支黑豆根内成虫数量减少,起到抗病效果。随后以抗性品种灰皮支黑豆根系总RNA为模板,利用RT-PCR技术获得了939 bp的Gm MIOX4基因的CDS序列,克隆得到质粒p GEM-MIOX4,再以得到的Gm MIOX4序列为材料构建过表达载体p CAM-MIOX4,为进一步研究Gm MIOX4基因功能奠定了基础。Two kinds of different resistance soybean（ Huipizhiheidou and Liaodou 15 ） were inoculated with SCN3 in the re- search. By real-time quantitative PCR method, GmMIOX4 gene expression changes were identified in early period and com- plete life cycle of SCN3. Real-time quantitative PCR results showed that： Under SCN3 infection, the relative expression of GmMIOX4 gene in roots of Liaodou 15 increased until 25 dpi after inoculation, the relative expression of GmMIOX4 gene in roots of Huipizhiheidou reached the highest level in 10 dpi, GmMIOX4 gene was induced and played an important role. Host＇ s nutrients was absorbed by SCN through syncytial to maintain normal growth and development period at 10 dpi, and this gene expression is induced, the gene can changes syncytial structure, inhibit nematode development, and thus played a dis- ease-resistant effect. Total RNA of Huipizhi Heidou was used as template, 939 bp CDS of GmMIOX4 genezone was obtained by RT-PCR. According to sequencing result, pGEM-MIOX4 over expression vector was cloned, it laid the foundation for further study of GmMIOX4 gene function.

关 键 词：GmMIOX4基因 大豆胞囊线虫 实时荧光定量PCR 克隆 过表达载体 

分 类 号：S435.651[农业科学—农业昆虫与害虫防治] Q943.2[农业科学—植物保护]