鳡胰岛素样生长因子1基因的克隆及其组织特异性表达分析  

Cloning and expression analysis of the insulin-like growth factor 1 in Elopichthys bambusa

作　　者：郁建锋[1] 李建林[2] 徐建荣[1] 卢祥云[1] 柏洁[1] 顾志良[1] 

机构地区：[1]常熟理工学院生物科学与工程系,江苏常熟215500 [2]中国水产科学研究院淡水渔业研究中心/农业部淡水鱼类遗传育种和养殖生物学重点开放实验室,江苏无锡214081

出　　处：《扬州大学学报（农业与生命科学版）》2012年第3期32-39,共8页Journal of Yangzhou University：Agricultural and Life Science Edition

基　　金：国家自然科学基金资助项目(31072025);教育部出国留学回国人员科研启动基金资助项目(2011);苏州市2010年度科技基础设施建设项目(SZSD201001);农业部淡水鱼类遗传育种和养殖生物学重点开放实验室开放基金资助项目(KY2009047)

摘　　要：为克隆鳡(Elopichthys bambusa)的胰岛素样生长因子1(IGF1)全长cDNA和启动子序列及明确其组织表达特征,采用反转录PCR(RT-PCR)、cDNA末端快速扩增(RACE)和染色体步移等技术,从鳡肌肉中获得了IGF1基因的全长cDNA(844bp)和上游启动子部分序列(627bp)。结果表明:IGF1基因包含218bp的5′端非翻译区、140bp的3′端非翻译区和486bp的开放性阅读框(ORF),编码161个氨基酸,包含前44个氨基酸残基为信号肽、70个氨基酸残基的4个功能结构域和C端47个氨基酸残基的延伸肽结构域。启动子无TATA框,但含有一成肌分化抗原(Myo D)结合位点。氨基酸同源性和系统发育分析发现,鳡IGF1与鲤形目其他鱼类的同源性较高(94.41%～99.38%),亲缘关系最近。半定量RT-PCR结果显示,IGF1在鳡的肝脏中表达最高,而心脏中未见其明显表达。该研究为明确IGF1参与鳡生长发育的机制研究及其在鳡的繁殖育种中的应用奠定了基础。The objective of the present research was to clone full length IGF1 cDNA sequence and to determine its expression in various tissues from Elopichthys bambusa. Reverse transcription PCR （RT-PCR）, rapid-amplification of cDNA ends （RACE） were used to amplify full-length of IGF1 cDNA. Semi-quantatative RT-PCR was used to compare IGF1 gene expression levels from various tissues. A full-length of IGF1 cDNA sequence in E. bambusa was obtained, which included a complete open reading frame （ORF） of 486 bp, a 5＇-untranslated region of 218 bp, and a 3＇-untranslated region of 140 bp. The putative peptide contained a signal sequence of 44 amino acid, a mature peptide of 70 amino acid in the functional domain, and an extended carboxyl-terminal peptide of 47 amino acid in domain E. IGF1 from E. bambusa showed an amino acid identity of 94.41%- 99.38% with that from Cypriniformes. The phylogenetic analyses showed that IGF1 from E. bambusa had closest relationship with other fishes in Cypriniformes. Furthermore, IGF1 mRNA levels were highest in liver but relatively low or absent in heart. A part of IGF1 promoter sequence was cloned by using Genome Walking, which included a Myogenic Differentiation Antigen （Myo D）. But there was not a consensus TATA box a bout 5＇ to the start site of transcription in IGF1. These results indicated the IGF1 promoter differed to the classic promoter. These findings provide clue for IGF1 involvement in the growth and development of E. bambusa and are potential basis for its effective breeding and propagation.

关 键 词：鳡 胰岛素样生长因子1 启动子 开放性阅读框 表达 

分 类 号：S917.4[农业科学—水产科学]