红螯螯虾网格蛋白轻链的克隆及制备抗体的检测  

Cloning of clathrin light chain of crayfish,Cherax quadricarinatus,and immunological specificity of antibodies

作　　者：黄家骏[1] 李钫[1] 杨丰[1] 

机构地区：[1]国家海洋局第三海洋研究所国家海洋局海洋生物遗传资源重点实验室,福建厦门361005

出　　处：《应用海洋学学报》2016年第2期205-210,共6页Journal of Applied Oceanography

基　　金：国家973课题资助项目(2012CB114401)

摘　　要：网格蛋白为三脚蛋白复合体,其介导的内吞作用是病毒侵染细胞的常用途径之一.而对虾白斑综合症病毒(white spot syndrome virus,WSSV)是对虾养殖业的主要病原,为了研究分析WSSV的侵染机制及进入宿主细胞的途径,在本研究中,克隆得到红螯螯虾(Cherax quadricarinatus)网格蛋白轻链的c DNA全长,并将其命名为Cq CLC(Genbank登录号为KR075008).进化树分析表明,其与黑腹果蝇(Drosophila melanogaster)的网格蛋白轻链进化距离较近.此外,在其蛋白序列中选择抗原性较好的片段合成多肽,并以此制备单克隆抗体,而后经免疫荧光和western blot检测从中筛选出具有特异性的抗体,为进一步对WSSV侵染虾细胞机制及入胞途径的研究奠定了基础.The clathrin triskelion is composed of 3 clathrin heavy chains and 3 light chains subunits,and clathrinmediated endocytosis is the way commonly infected by viruses. White spot syndrome virus（ WSSV） is a main pathogen in cultured shrimp,however,mechanism of WSSV infection is still unknown. In this study,completed sequence of clathrin light chain c DNA in crayfish,Cherax quadricarinatus,was obtained by cloning and RACE,designated Cq CLC（ Gen Bank accession number KR075008）. In phylogenetic tree analysis,evolutionary distance of clathrin light chain from the crayfish was closed to Drosophila melanogaster. Polypeptides of clathrin light chain were synthetized as antigens to immune the mouse. The immunological specificity of antibodies was validated by immunofluorescence and western blot. This work has provided an important foundation for further study of WSSV infection mechanism in shrimp cells.

关 键 词：海洋生物学 红螯螯虾 网格蛋白轻链 克隆 进化树分析 抗体特异性 

分 类 号：P735[天文地球—海洋生物学]