平颏海蛇毒鼠源单链抗体噬菌体展示文库的构建  

Construction of Phage Display Library for ScFv from Mouse Against Hydrophis curtus Venom

作　　者：杜宇 张皓冰 杨宇琪 汪妍 赵红燕[3] 林炽贤 DU Yu;ZHANG Haobing;YANG Yuqi;WANG Yan;ZHAO Hongyan;Lin Chixian(School of Fisheries and Life Science,Hainan Tropical Ocean University,Sanya Hainan 572022,China;Hainan Key Laboratory of Herpetological Research,Hainan Tropical Ocean University,Sanya Hainan 572022,China;College of Life and Environmental Sciences,Hangzhou Normal University,Hangzhou Zhejiang 311121,China)

机构地区：[1]海南热带海洋学院水产与生命学院,海南三亚572022 [2]海南热带海洋学院海南省两栖爬行动物研究重点实验室,海南三亚572022 [3]杭州师范大学生命与环境科学学院,杭州311121

出　　处：《海南热带海洋学院学报》2020年第5期23-28,共6页Journal of Hainan Tropical Ocean University

基　　金：海南省财政科技计划资助项目(ZDYF2018120,ZDKJ2016009-2-1);三亚市农业科技创新项目(2016NK13)。

摘　　要：为了研发新型小分子类抗海蛇毒药物,本研究利用平颏海蛇(Hydrophis curtus)毒免疫成年小鼠,使其产生免疫应答,以小鼠的脾脏总RNA为基础,反转录构建cDNA文库,分别扩增抗体轻链和重链可变区片段并通过重叠延伸PCR技术构建平颏海蛇毒小鼠单链抗体基因.构建完成后,该基因随后被成功重组至噬菌体,并利用大肠杆菌原核表达系统建立了单链抗体噬菌体展示文库.经多轮富集后,最终获得弱阳性克隆.本研究获得了一套构建平颏海蛇毒鼠源单链抗体噬菌体文库并筛选阳性克隆的方案.To develop a novel antibody with small molecular mass against sea snake venom,we constructed the mouse single-chain antibody variable fragment(ScFv)by SOE-PCR based on the light and heavy chain variable region fragments,which were respectively amplified from the cDNA library corresponding to the total spleen RNA from the mouse immunized by H.curtus venom.The ScFv gene was then successfully ligated to the phage genome,followed by the establishment of a ScFv library in E.coli.After several cycles of enrichment,weak positive clones were detected from the ScFv library.In the present study,a phage display library of mouse ScFv against H.curtus venom was constructed and the positive clones were screened.

关 键 词：平颏海蛇(Hydrophis curtus) 蛇毒 单链抗体 基因工程 噬菌体展示技术 

分 类 号：Q178.53[生物学—水生生物学] Q785[生物学—普通生物学]