龙眼DlWRKY57基因克隆与拟南芥遗传转化  

Cloning and Arabidopsis Genetic Transformation of DlWRKY57 Gene in Dimocarpus longan

作　　者：黄幸[1] 丁峰[2] 潘介春[1] 张树伟[3] 徐炯志[1] 王金英 李琳 王颖 李浩然 刘一 Huang Xing;Ding Feng;Pan Jiechun;Zhang Shuwei;Xu Jiongzhi;Wang Jinying;Li Lin;Wang Ying;Li Haoran;Liu Yi(College of Agriculture,Guangxi University,Nanning,530004;Guangxi Crop Genetic Improvement and Biotechnology Laboratory,Guangxi Academy of Agricultural Science,Nanning,530004;Institute of Horticulture,Guangxi Academy of Agricultural Science,Nanning,530007)

机构地区：[1]广西大学农学院,南宁530004 [2]广西壮族自治区农业科学院,广西作物遗传改良生物技术重点开放实验室,南宁530004 [3]广西壮族自治区农业科学院园艺研究所,南宁530007

出　　处：《分子植物育种》2021年第1期149-157,共9页Molecular Plant Breeding

基　　金：国家荔枝龙眼产业技术体系(CARS-33-10);国家现代农业产业技术体系广西荔枝龙眼创新团队(nycytxgxcxtd-02)共同资助。

摘　　要：以龙眼成熟叶片为材料,利用实验室构建的龙眼转录组数据库,筛选出Dl WRKY57基因c DNA序列,对其开放阅读框(ORF)进行克隆和生物信息学分析。结果表明,Dl WRKY57基因ORF长度为894 bp,编码297个氨基酸,此蛋白属于WRKY基因家族中的域a类,存在1个WRKY结合位点。通过生物信息学分析表明,该蛋白属于非跨膜亲水蛋白,预测定位于细胞核,存在45个氨基酸磷酸化位点。其二级结构由无规则卷曲、琢-螺旋和延伸链构成,其中无规则卷曲为二级结构中最主要的结构元件,并且二级结构与三级结构预测结果高度一致。同源基因进化树分析表明该基因与木薯亲缘关系最近。以p MD18-T和p BI121载体为基础,成功构建了植物超表达载体p BI121-Dl WRKY57,利用农杆菌花序法转化拟南芥,经PCR分子检测,获得含Dl WRKY57基因的拟南芥植株,该超表达载体的遗传转化为Dl WRKY57基因功能的深入研究提供依据。The mature leaves of longan were used as material.The Dl WRKY57 gene c DNA sequence was screened from longan transcriptome database of Dimocarpus longan constructed in the laboratory,and its open reading frame(ORF)was used for cloning and bioinformatics analysis.The results showed that the Dl WRKY57 ORF sequence was894 bp in length,encoding 297 amino acids,a member of classⅡa and there is a WRKY binding site.Bioinformatics analysis showed that the protein was a non-transmembrane hydrophobic protein,which was predicted to be located in the nucleus,with 45 amino acid phosphorylation sites and 1 WRKY binding site.The secondary structure consisted of random coil,alpha-helix and extended chain.Random crimp is the most important structural element in the secondary structure,and the prediction results were highly consistent between secondary structure and tertiarystructure.The analysis of protein phylogenetic tree indicated that Dl WRKY57 gene was the closest relative to homologous gene of Manihot esculenta.The plant overexpression vector p BI121-Dl WRKY57 was successfully constructed based onp MD18-T vectorand p BI121,andtransferred into Arabidopsis thalianabyfloral dip.Transgenic plants with Dl WRKY57 gene were obtained successfully after PCR molecular detection.The successful genetic transformation of the overexpression vector will provide a basis for the further study of Dl WRKY57 gene function.

关 键 词：龙眼(Dimocarpus longan) DlWRKY57 基因克隆 超表达载体构建 

分 类 号：S667.2[农业科学—果树学]