油菜沪油15中AP2/ERF-B3亚族转录因子的克隆和生物信息学分析  

CLONING AND BIOINFORMATIC ANALYZING OF TRANSCRIPTION FACTOR AP2/ERF-B3 SUBFAMILY GENES FROM BRASSICA NAPUS L. HUYOU 15

作　　者：庄静[1] 周熙荣[2] 孙超才[2] 管帮超[3] 彭日荷[4] 乔玉山[1] 章镇[1] 熊爱生[4] 姚泉洪[4] 

机构地区：[1]南京农业大学园艺学院 [2]上海市农业科学院作物育种栽培研究所,上海201106 [3]上海市崇明县农业技术推广中心,上海202150 [4]上海市农业科学院生物技术研究所,上海201106

出　　处：《分子细胞生物学报》2008年第3期192-206,共15页Journal of Molecular Cell Biology

基　　金：863项目(2006AA10Z117;2006AA06Z358;2008AA10Z401);上海市科技启明星和优秀学科带头人计划~~

摘　　要：AP2/ERF转录因子家族广泛存在于植物中,参与植物细胞周期、生长发育以及生物和非生物胁迫相关基因表达调控。本文利用油菜UniGene数据库,以拟南芥AP2/ERF-B3亚族转录因子保守序列为信息探针,分离得到2个油菜AP2/ERF-B3亚族的转录因子BnaERFB3-1和BnaERFB3-2。通过PCR和RT-PCR方法分别从双低甘蓝型油菜沪油15的DNA和cDNA中克隆了上述基因。序列分析显示,克隆的BnaERFB3-1-Hy15和BnaERFB3-2-Hy15转录因子与电子克隆的基因序列差异很小,均只有1个氨基酸位点不同,且都没有内含子。从氨基酸序列的相似性、组成成分、理化性质、疏水性/亲水性、序列比对、进化树、功能域、二级结构、三级结构、无序化特性等方面进行了预测和较为全面的分析。结果显示BnaERFB3-1-Hy15和BnaERFB3-2-Hy15是亲水性蛋白,在蛋白质的三级结构上与AtERF5相似,BnaERFB3-1-Hy15和BnaERFB3-2-Hy15蛋白无序化程度大于拟南芥AtERF5。通过分析EST丰度显示,BnaERFB3-1的表达集中在种子中,而BnaERFB3-2的表达则集中在根中。另外,将上述基因分别构建入酵母表达载体和植物双元表达载体,为深入研究该基因在油菜抗逆调控中的作用奠定了基础。ABSTRACT AP2/ERF is a large family of transcription factors in plant. Genes in the AP2/ERF family encode transcriptional regulators with a variety of functions involved in the developmental and physiological processes in plants. Two AP2/ERF family transcriptional regulators （BnaERFB3-1 and BnaERFB3-2） were isolated from B. napus by in silico cloning method using the conserved domain amino acid sequence of A. thaliana AP2/ERF-B3 subfamily as probe. Based on the sequences of BnaERFB3-1 and BnaERFB3-2, we isolated the BnaERFB3-1-Hy15 gene and BnaERFB3-2-Hy15 gene from winter and spring type B. napus L. cv Huyou15 by RT-PCR and PCR using eDNA and DNA as template. DNA sequencing and analyzing indicated that there was only one amino acid residue difference between BnaERFB3-1 and BnaERFB3-1-Hy15, BnaERFB3- 2 and BnaERFB3-2-Hy15, respectively. No intron localized on the two genes from Huyou15. Then, deduced amino acid sequence, composition, hydrophobicty and hydrophilicity, physical and chemical characterization, phylogenetic tree, conserved domain sequences, function domain, molecular modeling, and folding state were predicted and analyzed. BnaERFB3-1-Hy15 and BnaERFB3-2-Hy15 were hydrophilic protein. The two proteins and AtERF5 have similar three-dimension structure. The disordered residues of protein BnaERFB3-1-Hy15 and BnaERFB3-2-Hy15 were higher than that of AtERF5. BnaERFB3-1 was mainly expressed in seed, while BnaERFB3-2 was mainly expressed in root. Moreover, those genes were successfully constructed into the recombinant plasmids of plant expression vector and yeast expression vector, which established a base for transformation of oilseed and studies of those genes function in abiotic stresses of B. napus.

关 键 词：转录因子AP2/ERF—B3亚族 生物信息学 进化树 油菜沪油15 

分 类 号：Q943.2[生物学—植物学] S565.4[农业科学—作物学]