陆地棉(Gossypium hirsutum L.)GhWRKY75的克隆及表达载体构建  

Cloning and expr ession vector constr uction of GhWRKY75 in gossypium hir sutum L.

作　　者：张鹏飞[1] 刘戈辉 郭文婷[1] 张薇[1] Zhang Pengfei;Liu Gehui;Guo Wenting;Zhang Wei(Key Laboratory of Xinjiang Oasis Agriculture /College of Agronomy,Shihezi University,Shihezi,Xinjiang 832003,China)

机构地区：[1]新疆绿州农业重点实验室/石河子大学农学院,新疆石河子832003

出　　处：《石河子大学学报（自然科学版）》2018年第3期330-336,共7页Journal of Shihezi University(Natural Science)

基　　金：国家自然科学基金项目(31260358)

摘　　要：目的从棉花抗病品种中克隆WKKY转录因子基因,为棉花抗枯萎病分子育种提供重要的基因资源。方法以枯萎病菌诱导后棉花基因表达谱中差异表达的WRKY基因片段为探针序列,以陆地棉的抗枯萎病品种'中棉所12'的根部c DNA为模板。运用电子拼接及RT-PCR技术,克隆得到一个新的基因,将该基因命名为GhWRKY75(登录号:MH138002)。结果序列分析发现,GhWRKY75基因开放阅读框全长为513bp,编码一个含170个氨基酸残基的蛋白,具有1个WRKY结构域及1个典型的C2H2型锌指结构,属于第Ⅱ类WRKY转录因子家族。该基因编码蛋白分子量为19.51kd,等电点为9.30,脂肪指数为60.12,属于疏水性蛋白,基因的蛋白二级结构由α螺旋、延伸链和无规则卷曲组成。同源基因进化树分析表明该基因与雷蒙德氏棉亲缘关系最近。结论:从棉花中克隆得到一个新的GhWRKY75基因,构建了该基因的植物过表达载体,为进一步研究该基因功能奠定了基础。Objective The anim of the study was to clone the WRKY transcription factor gene fom the cotton resistent caltivars and frovide important penetic resources for mdecular breading of resistance to Fusaricm wilt. R esults A new gene of WRKY was cloned used the c DNA of the roots of ＇Zhongmiansuo12＇ as template by using RT-PCR. This was named GhWRKY75（Gen Bank accession number： MH138002）. Sequence analysis showed that the open reading frame of GhWRKY75 gene was 513 bp, and encoded a protein of 170 amino acids. The protein contained a WRKY domain and a typical C2H2 zinc finger structure, which indicated this gene belongs to the II type WRKY family transcription factor. The molecular weight of GhWRKY75 is 19.51 k D,isoelectric point is 9.30, fat index is 60.12, and is a hydrophobic protein. The secondary structure of GhWRKY75 protein consists of α-helix, extended chain and random coil. Phylogenetic tree analysis o showed that GhWRKY75 is closely related to homologous gene of Gossypium raimondii. Conclusion The successful cloning of a new gene of GhWRKY75 from the cotton and construction of the plant overexpression vector GhWRKY75 gene.This study laid a foundation for further study on the function of GhWRKY75.

关 键 词：陆地棉 基因克隆 WRKY转录因子 载体构建 

分 类 号：S562[农业科学—作物学]