大豆MYB转录因子GmMYBZ2的鉴定、突变分析及原核表达  

Characterization and Mutation of Soybean MYB Transcription Factor GmMYBZ2 and Its Prokaryotic Expression

作　　者：杜海[1,2] 刘蕾[1,2] 唐晓凤[1,2] 赵艳丽[2] 高杰[2] 吴燕民[2] 黄玉碧[1] 唐益雄[2] 

机构地区：[1]四川农业大学玉米研究所,雅安625014 [2]中国农业科学院生物技术研究所,北京100081

出　　处：《农业生物技术学报》2009年第2期301-306,共6页Journal of Agricultural Biotechnology

基　　金：中央级公益性科研院所基础科研业务费专项资金(No.基本2008-17)资助

摘　　要：大豆(Glycine max)GmMYBZ2基因是典型的植物R2R3-MYB转录因子家族成员之一。通过DNAMAN和酵母(Saccharomyces cerevisiae)单杂交系统分别对其蛋白结构和转录活性进行了分析鉴定,并在大肠杆菌(Escherichia coli)中进行了原核表达。GmMYBZ2除含有典型的R2R3-MYB转录因子的结构特征外,在C端含有1个少有的保守氨基酸基序PDLNLELTIS及一个隐藏的锌指结构。基因组序列分析表明,在263～395bp位含1个132bp的内含子。为明确GmMYBZ2C端保守氨基酸基序及锌指结构对目的蛋白转录活性的影响,分别进行了PCR介导的序列删除突变。酵母单杂交系统分析显示,内含子、保守基序及锌指结构对目的蛋白的转录活性均有抑制作用;原核表达显示,目的蛋白能在补充有稀有密码子的大肠杆菌中成功表达。Soybean （Glycine max） GmMYBZ2 is one of typical plant R2R3-MYB family genes. In this study, its protein characterization and transcriptional activity were analyzed by DNAMAN and yeast （Saccharomyces cerevisiae） one-hybrid system, respectively. Moreover, the prokaryotic expression vector was constructed and expressed in Escherichia coli as well. Results showed that GmMYBZ2 not only exhibited typical features of the R2R3-MYB transcription factors, but also had a conserved amino acid motif and a predicted zinc finger region at its C-termini. Genomic DNA sequence analysis showed that it contained an intron in the R3 domain which was located at 263-395 bp. In order to investigate the infections of these motifs in GmMYBZ2＇s transcriptional activity, the conserved motif and zinc finger region were detected by PCR. The effect of the altered proteins were monitored using yeast one-hybridize, which showed that GmMYBZ2 had transcriptional activation function. And the intron, conserved motif and zinc finger region might repress the transcriptional activity of GmMYBZ2. Moreover, the encoding sequence of GmMYBZ2 gene was inserted into pET30a to construct prokaryotic expression vector pET-ZD2 which was then transformed into E. coli BL21 （DE3） pLysS and Rosetta2 （DE3） pLysS, respectively. Results showed that GmMYBZ2 could be expressed in E. coli Rosetta2 （DE3） which contained complementary rare codons successfully.

关 键 词：MYB转录因子 突变 酵母单杂交 原核表达 

分 类 号：S188[农业科学—农业基础科学]