鲢转铁蛋白基因cDNA的克隆及其在胚胎期的表达分析

Whole sequence of cDNA cloning and tissue expression analysis of transferrin gene in Hypophthalmichthys molitrix during embryogenesis

克隆鲢转铁蛋白（transferrin，Tf）基因的全长cDNA序列，并对鲢转铁蛋白的组织表达模式、胚胎发育过程中的时序表达模式进行分析。结果显示，该cDNA全长2365bp，包含2025bp开放阅读框（ORF），编码674个氨基酸。鲢转铁蛋白与草鱼转铁蛋白的同源性高达74％，与人乳铁蛋白同源性最低，为39％，与其他鲤科鱼类转铁蛋白的同源性约65％～73％，与非鲤科鱼类的同源性约43％～50％。系统进化树分析显示，鲢转铁蛋白基因与斑马鱼、草鱼、鲤、鲫等几种鲤科鱼类亲缘关系最近，单独聚为一支。鲢转铁蛋白基因仅在肝脏和脾脏中表达（肝脏中表达量高于脾脏）。在胚胎发育过程中，转铁蛋白基因对原肠中期后的器官分化和形态建成起到了一定的作用，而未影响原肠中期前的细胞分裂增殖。

Full length cDNA of silver carp transferrin, Hypophthalmichthys molitrix, was first cloned using reverse transcription polymerase chain reaction （RT-PCR） and SMART RACE methods. The entire transferrin eDNA sequence is 2 365 bp long and the open reading frame is 2 025 bp and en- codes a protein with 674 amino acids. Molecular characteristics of the gene were forecasted by online molecular softwares. The results showed that it is composed of two domains,with a signal peptide of 15 amino acids which located at the 21st amino acid of N-terminal. Silver carp transferrin gene has high homology with the other species, sharing the highest identity of 74 % with grass carp Ctenopharyngodon idella. Phylogenetic tree analysis revealed that transferrin of five Cyprinid fish,Danio rerio, Carassius auratus ,Cyprinus carpio , Ctenopharyngodon idella and Hypophthalmichth ys rnolitrix were classified together. Siler carp transferrin mRNA only expressed in liver and spleen （a higher expression level in liver）. During embryogenesis, the expression of transferring mRNAs was first detected at the gastrula stage and the expression level increased steadily until the tail bud embryo. Transferrin mRNA showed marked reduction after the stage of tail bud embryo until larvae. Results from the present study indicated that transferrin played an important role in organogenesis during embryogenesis.