当归延伸因子AsEF-1β基因的克隆及胁迫应答分析

Cloning and Analysis of Response to Stress of Elongation Factor 1β Gene from Angelica sinensis

延伸因子1β(EF-1β)是蛋白质生物合成过程中肽链延长必需的调节因子之一。该研究采用同源克隆和RACE扩增技术克隆当归EF-1β基因序列,分析该基因序列特征、蛋白结构特点及UV-B辐射胁迫下的组织响应表达,以揭示当归栽培生境变迁过程中对UV-B胁迫适应的分子机制。结果显示:(1)成功克隆获得当归EF-1β基因全长序列(950 bp),编码225个氨基酸,命名为AsEF-1β(GenBank登录号:MG736314);AsEF-1β蛋白的分子量为24.5 kD,理论等电点为4.48,属亲水性氨基酸,在其C末端具有一个EF-1B超蛋白家族的典型结构域和保守区,鸟嘌呤核苷酸交换结构域;其氨基酸序列与同为伞形科的胡萝卜氨基酸序列相似性最高,达93%。(2)qRT-PCR分析结果显示,AsEF-1β基因在当归根部的表达量显著高于茎和叶(P<0.05);UV-B辐射胁迫下,茎及叶中的表达量均上调,分别是自然光照处理的2.43和3.76倍。研究表明,AsEF-1β基因可能参与当归对UV-B辐射胁迫的适应过程,为深入研究其在药用植物生长发育、逆境抗性形成及药效物质的生物合成代谢过程的生态调控奠定了基础。

Elongation factor 1β (EF-1β) is one of the essential regulatory factors of peptide chain prolongation in the process of protein biosynthesis.This study cloned the EF-1β gene sequence from Angelica sinensis using homologous cloning and RACE amplification techniques.We analyzed the sequence characterization,protein structure characteristics and expression profiling of EF-1β gene under UV-B radiation stress,to reveal the molecular mechanism of adaptation to UV-B radiation stress in the process of cultivation habitat change of A.sinensis.The results showed that:(1)the full-length sequence of EF-1β gene(950 bp),encodes 225 amino acids,which was named AsEF-1β gene(GenBank accession number:MG736314).The molecular weight of AsEF-1β is 24.5 kD,theoretical isoelectric point of 4.48.AsEF-1β belongs to hydrophilic amino acid,with the typical domain and conservative domain of family of EF-1B super protein,and has a guanine nucleotide exchange domain at its C-terminal.The amino acid sequence of AsEF-1β gene showed the highest similarity with that of Daucus carota L.var.sativa Hoffm.,which belongs to the same family of Umbrella.(2)the qRT-PCR analysis results indicated that AsEF-1β gene in root of A.sinensis was significantly higher than that in stem and leaf(P<0.05).Under UV-B radiation stress,the relative expression level of stem and leaf were up-regulated,which was 2.43 times and 3.76 times that of natural light condition(P<0.05),respectively.Studies showed that AsEF-1β gene may be involved in the adaptation process of A.sinensis to UV-B radiation stress,which will be laid for further exploration on its ecological regulations in the growth and development,formation of stress resistance and biosynthesis of pharmacodynamic substances of medicinal plants.