鲢鱼恒定链cDNA的克隆及其分子结构分析  

cDNA cloning and molecular structure analysis ofAristichthys molitrix Invariant chain

作　　者：叶显峰[1] 孟凡涛[1] 刘洪明 祖国掌[1] 

机构地区：[1]安徽农业大学安徽省人兽共患病重点实验室,合肥230036 [2]烟台市动物卫生监督所,山东烟台264000

出　　处：《淡水渔业》2013年第3期9-14,共6页Freshwater Fisheries

基　　金：国家自然科学基金(No.31172306)

摘　　要：为研究鱼类恒定链(Invariant chain,Ii)的结构与功能,应用RT-PCR和RACE(rapid amplification of cDNAends)法,从脾脏细胞中克隆了鲢(Aristichthys molitrix)Ii基因cDNA,并进行了测序。该基因cDNA全长为1 136bp,其中开放阅读框为699 bp,编码232个氨基酸。比对氨基酸序列发现,鲢与其他物种的Ii链有相同的结构域。将Ii基因cDNA片段和表达不同结构域的DNA片段分别插入真核表达质粒pEGFP-C1,转染真核细胞系COS7,发现鲢Ii跨膜区和胞浆区在细胞内具有定位作用。根据推测的氨基酸序列,分析不同种间Ii的同源性,表明鲢与斑马鱼(Brachydanio rerio)的同源性最高(78%),与人等哺乳动物Ii的同源性较低(30%～40%)。进一步模拟构建和比较不同物种Ii链的3D结构,显示链鱼和小鼠、人和鸡的Ii链结构非常相似,甚至在一些具有重要作用的氨基酸是相同的。上述结果表明,作为重要的免疫分子,不同脊椎动物的Ii链不仅在遗传上具有高度同源性,而且在结构方面仍保持着高度的相似性。To know the structure and function of fish Invariant chain（Ii）,we cloned and sequenced Aristichthys molitrix Ii gene cDNA fromsleen cells by RT-PCR and RACE.The full length Ii-cDNA was 1∶136 bp,its open reading frame（ORF） was 699bp encoding 232 amino acids.Sequence alignment showed that the Ii had the same structure domains as other species.The li gene and its functional segments domains were cloned into an eukaryocyte expression vector pEGFP-C1,respectively,and further these reconstructed vectors were transfected into eukaryotic expression cell COS7.The result showed that A.molitrix Ii was located on plasma membrane as a transmembrane glycoprotein.The deduced amino acid sequences showed that the A.molitrix had a similarity of 78% to Brachydanio rerio and had a lower similarity of 30%~40% to human and other mammals.The Ii 3D molecular structure of different species was simulated and constructed.The comparison of them revealed a similarity in the structure among Iis of A.molitrix,mouse,human and chicken,especially some amino acid residues at the important sites.These results demonstrated that Ii as an important immune molecule had a high degree of homology in genetics and a similarity in structure among different vertebrates.

关 键 词：恒定链 鲢(Aristichthys molitrix) RACE 克隆 定位分析 

分 类 号：S917.4[农业科学—水产科学]