牙鲆胰岛素样生长因子结合蛋白IGFBP-1 cDNA全长的克隆及表达分析

利用RACE-PCR技术,从牙鲆(Paralichthys olivaceus)的肝脏组织总RNA中克隆得到胰岛素样生长因子结合蛋白-1(insulin-like growth factor binding protein 1, IGFBP-1)基因的全长cDNA序列,该cDNA全长为1070 bp,开放阅读框为729 bp,编码242个氨基酸。通过系统进化树分析,牙鲆IGFBP-1与鱼类IGFBP-1基因聚为一支；通过同源性比对,牙鲆IGFBP-1基因的核苷酸序列与大菱鲆同源性最高,为95%,而其推导的氨基酸序列与其它鱼类如大菱鲆、五条鰤、黄金鲈、红点鲑、鲤鱼和斑马鱼的同源性分别为89%、89%、84%、79%、67%和67%。半定量RT-PCR分析表明,牙鲆IGFBP-1基因存在母源转录本,合子基因在孵化前的胚胎阶段及早期仔鱼中仅有较低水平的表达,在后期仔鱼中表达逐渐增高；牙鲆IGFBP-1基因在肝脏中表达量最高,在胃、脾、肠、性腺、肾、鳃、脑、心脏和肌肉中也有不同程度的表达。

The full-length cDNA cloning and gene expression analysis of insulin-like growth factor binding protein 1 (IGFBP-1)in Japanese flounder ( Paralichthys olivaceus )

The full-length cDNA of insulin-like growth factor binding protein 1 gene(IGFBP-1) was obtained from the liver of Japanese flounder(Paralichthys olivaceus)using RACE method. The complete cDNA of P.olivaceus IGFBP-1 is 1 070 bp and its ORF includes 729 bp encoding 242 amino acid residues.The IGFBP-1 was highly homologous with IGFBP-1 gene in fish through phylogency analysis. The nucleotide sequence of IGFBP-1 from P.olivaceusshared 95% homology with that of Scophthalmus maximu,and the deduced amino acid analysis showed that the sequence similarities betweenP.olivaceusand S.maximu,Seriola quinqueradiata,Perca flavescens,Salvelinus alpines,Cyprinus carpio andDanio rerio,are 89%,89%,84%,79%,67% and 67%,respectively. Semi-quantitative RT-PCR results demonstrated that IGFBP-1 mRNA was detected in unfertilized egg,fertilized egg,embryonic and larval development,and various adult tissues.Moreover,real-time quantitative RT-PCR results showed that IGFBP-1 gene had the highest expression in adult liver and was expressed in a stage-specific manner during embryonic and larval development.