文献类型：期刊文献

中文题名：当归MYB4转录因子基因的克隆及表达分析

英文题名：Cloning and Expression Analysis of Transcription Factor MYB4 Gene from Angelica sinensis

作者：杨雪[1];雒军[2];杨彩霞[1];王振恒[1];夏琦[2];王引权[1]

第一作者：杨雪

机构：[1]甘肃中医药大学药学院;[2]甘肃中医药大学科研实验中心

第一机构：甘肃中医药大学药学院（西北中藏药协同创新中心办公室）

年份：2018

卷号：47

期号：12

起止页码：48

中文期刊名：河南农业科学

外文期刊名：Journal of Henan Agricultural Sciences

收录：CSTPCD;;北大核心:【北大核心2017】；CSCD:【CSCD_E2017_2018】；

基金：国家重点研发计划专项(2017YFC1700705);国家自然科学基金项目(81660625);甘肃省高校协同创新科技团队支持计划项目(2016C-05FF09)

语种：中文

中文关键词：当归;阿魏酸合成;MYB转录因子;基因克隆;表达分析

外文关键词：Angelica sinensis;Ferulic acid biosynthesis;MYB transcription factor;Gene cloning;Expression analysis

摘要：为探索当归(Angelica sinensis)药效成分阿魏酸生物合成中MYB蛋白的结构和功能,从当归中克隆转录因子基因MYB4,并进行序列分析和表达分析。根据MYB4转录因子的DNA保守结构域,结合同源克隆及RACE的原理,运用RT-PCR技术从当归中克隆MYB4基因cDNA的全长序列;利用生物信息学分析工具对该基因编码蛋白质的基本性质、结构特点及生物学功能进行初步预测;应用qRT-PCR分析该基因的表达特征;使用重组技术组装原核表达载体p GEX-4T-3-As MYB4,热击法转化至大肠杆菌BL21(DE3),IPTG(异丙基硫代-β-D-半乳糖苷)诱导重组蛋白表达。结果表明,成功克隆1个MYB4基因,命名为As MYB4,在GenBank中注册,登录号为MG736315;序列分析表明,As MYB4的cDNA全长为1 189 bp,包括804 bp的开放阅读框,编码267个氨基酸,为典型的R2R3-MYB类转录因子家族成员; As MYB4与胡萝卜MYB308的同源性达到93%,且亲缘关系最近。组织特异性表达分析表明,As MYB4基因在当归叶片中的转录水平最高,分别是叶柄、根的1. 1、2. 0倍;异源表达分析表明,As MYB4蛋白以包涵体形式存在,分子质量为30 ku。
To explore the structure and function of MYB protein involved in the biosynthesis of ferulic acid in Angelica sinensis,the transcription factor gene of MYB4was rapidly isolated and cloned from Angelica sinensis,and sequence analysis and expression analysis of MYB4gene were performed.According to the DNA conserved domain of MYB4transcription factor,combined with the principle of homologous cloning and RACE,the full-length cDNA of MYB4gene was cloned from Angelica sinensis by RT-PCR;The basic properties,structural characteristics and biological functions of the protein encoded by MYB4gene were preliminarily predicted by using bioinformatics analysis tools;The expression characteristics of MYB4gene were analyzed by qRT-PCR;Recombination technology was used to assemble prokaryotic expression vector pGEX-4T-3-AsMYB4,which was transformed into Escherichia coli BL21(DE3)by heat shock,and IPTG was used to induce the expression of recombinant protein.The results showed that a MYB4gene was cloned and named AsMYB4,which was registered in GenBank and the accession number was MG736315;Sequence analysis showed that the full-length cDNA of AsMYB4gene was1189bp,including a804bp open reading frame,and encoding267amino acids.It was a typical member of R2R3-MYB transcription factor family;The homology between AsMYB4and Daucus carota MYB308reached93%,and AsMYB4had the closest relationship with Daucus carota MYB308.Tissue specific expression analysis showed that transcription level of AsMYB4gene was highest in leaves of Angelica sinensis,which was1.1and2.0times of petioles and roots,respectively;Prokaryotic expression analysis showed that AsMYB4protein existed as inclusion bodies with a molecular mass of30ku.