河川沙塘鳢GH基因及侧翼的克隆与生物信息学分析

Cloning and bioinformatics analysis of GH gene and its flanking region in Odontobutis potamophila

利用长片段PCR、基因组步移法及T-A克隆技术,成功克隆出河川沙塘鳢( Odontobutis potamophila ) GH 基因全长序列,提交至GenBank获得登录号MH717101。河川沙塘鳢 GH 基因全序列长5120 bp,5′端和3′端侧翼序列长度分别为920、682 bp,经预测,上游区域含有MF3、GAGA factor、HNF-3alpha、C/EBP、R2、GATA-1等多种转录调控元件;转录单元长3 518 bp,包含4个内含子和5个外显子,4个内含子大小分别为75、333、2 070和121 bp,5个外显子长度分别为150、131、117、138和373 bp,第3内含子长达2 070 bp,其长度是翘嘴鲌( Culter alburnus )第3内含子的4倍以上;开放阅读框区(open reading frame,ORF)为594 bp,编码由197个氨基酸残基组成的蛋白质多肽。河川沙塘鳢与鲈形目、鲉形目、鲤形目、鲇形目等13种鱼类的氨基酸序列同源性比较表明,其与彼氏冰鰕虎鱼( Leucopsarion petersii )同源性最高,为83.8%,与草鱼( Ctenopharyngodon idella )同源性最低,为49.1%,在180~197氨基酸序列处,河川沙塘鳢表现出高度不保守性,与所列鱼类氨基酸排列均完全不一致。该研究结果为进一步研究河川沙塘鳢 GH 基因的表达、功能及其转录调控特征奠定了分子基础。

The full-length sequence of the Odontobutis potamophila GH gene was successfully cloned by using long segment PCR,genome walking method and T-A cloning technology,which was submitted to GenBank to obtain the login number MH717101. The full-length of the GH gene of Odontobutis potamophila is 5120 bp,the length of the 5′-end and3′-end lateral sequence is 920 bp and 682 bp. It is predicted that the upstream region contains MF3,GAGA factor,hnf-3alpha,C/EBP,R2,GATA-1 and other transcriptional regulatory elements,respectively. The transcription unit is3 518 bp long and contains 4 introns and 5 exons. 4 introns are 75,333,2 070 and121 bp;5 exons are150,131,117,138 and373 bp, respectively. The third intron is2 070 bp,the length was more than 4 times of the culter third intron. The open reading frame area (ORF) is 594 bp,encoding a protein polypeptide composed of197 amino acid residues. Comparison of amino acid sequence homology between Odontobutis potamophila and Perciformes,Scorpaeniformes,Cypriniformes,Siluriformes with13 fishes showed that the homology was the highest to Leucopsarion petersii, at 83.8%,and the homology to Ctenopharyngodon idella was the lowest,at 49.1%,the Odontobutis potamophila to show highly nonconservative at180-197 amino acid sequence and the amino acid arrangement was completely different with the listed fish. The results of this study laid a molecular foundation for further study on expression,function and transcriptional regulation of GH gene in Odontobutis potamophila.