日本白鲫IGF-1基因全长cDNA克隆及组织表达分析

The Cloning of IGF-1 cDNA and Tissue Distribution in Japanese Crucian Carp(Carassius cuvieri)

通过已获得的多个物种IGF-1基因的cDNA序列,设计合成简并引物,用日本白鲫的肝脏总RNA反转录获得的cDNA做模板,克隆获得了IGF-1中间序列.在此基础上,通过SMART RACE,获得了IGF-1全长cDNA序列.经过序列对比分析,所得到的序列是日本白鲫(Carassius cuvieri)IGF-1 cDNA全长,其中开放阅读框为486 bp,编码含161个氨基酸的蛋白质,该蛋白质含有保守的信号肽、B、C、A、D和E结构域和6个半胱氨酸残基.同时,系统进化分析表明,在进化过程中IGF-1基因在鱼类中保持着高度保守的进化特征.IGF-1在日本白鲫中广泛表达于多个组织,尤其在肝脏中表达量最高.在垂体、心脏、肾脏和肌肉中有较高的表达.而在脑、脾脏、精巢和卵巢中的表达量较低.

In this investigation,we cloned the partial cDNA of IGF-1 in Japanese crucian carp by using a pair of degenerate primers based on the conservative sequence of obtained sequences from more species.The full length cDNA was then obtained by SMART RACE,which contained an ORF of 486 bp that encoded a protein of 161 amino acid residues containing signal peptide,B,C,A,D and E domains and six cysteine residues.Sequence alignment and phylogenetic analysis revealed that IGF-1 of fishes is conservative in evolu-tion.The study on tissue distribution indicated IGF-1 of Japanese crucian carp had remarkably similar ex-pression patterns in adult body tissues.The highest level of the expression occurred in liver.The higher lev-el of the expression appeared in pituitary,heart,kidney and muscle,while the lower level of the expression occurred in brain,spleen,testis and ovary.