创伤弧菌溶细胞素噬菌体单链抗体库的构建  

Construction of mouse- derived phage display library of scFv fragments against Vibrio vulnificus VvhA

作　　者：潘振 陈杰敏 胡成进[2] Zhen Pan;Jiemin Chen;Chengjin Hu(Department of Clinical Laboratory,The Affiliated Changzhou No.2 People’s Hospital of Nanjing Medical University,Changzhou 213000,China;Department of Laboratory Medicine,960 Hospital of PLA,Jinan 250031,China)

机构地区：[1]南京医科大学附属常州第二人民医院检验科,江苏常州213000 [2]中国人民解放军第960医院实验诊断科,山东济南250031

出　　处：《中华卫生应急电子杂志》2021年第5期287-291,共5页Chinese Journal of Hygiene Rescue(Electronic Edition)

基　　金：全军医学科技“十二五”重点项目(BWS12J014)。

摘　　要：目的为获得高特异性创伤弧菌溶细胞素单链抗体,利用噬菌体展示技术建立鼠源性噬菌体单链抗体库。方法用溶细胞素蛋白免疫BALB/c小鼠后,取脾脏提取总RNA,逆转录获得cDNA。以cDNA为模板扩增出抗体的轻链和重链可变区基因片段,通过OE-PCR技术拼接成完整scFv基因片段,构建重组噬菌粒pComb3XSS-scFv,电转化到大肠杆菌XL1-Blue构建噬菌体单链抗体库。结果构建鼠源性的噬菌体单链抗体文库其重组率为78%,库容量为3×10^(8)。结论成功构建鼠源单链抗体噬菌体文库,为创伤弧菌感染的快速诊断奠定基础。Objective To obtain highly specific VvhA cytolysin single-chain antibody,we used phage display technology to establish a murine phage single-chain antibody library.Methods After immunizing BALB/c mice with cytolysin protein,the total RNA was extracted from the spleen,and cDNA was obtained by reverse transcription.The cDNA was used as a template to amplify the antibody light chain and heavy chain variable region gene fragments,and OE-PCR technology was used to splice a complete scFv gene fragment.The recombinant phagemid pComb3 XSS-scFv was constructed and electrotransformed into E.coli XL1-Blue to construct a phage single-chain antibody library.Results The recombination rate of the mouse-derived single-chain antibody phage library was 78% and the library capacity was 3×10^(8).Conclusion The murine single-chain antibody phage library is successfully constructed and lays the foundation for the early diagnosis of Vibrio vulnificus infection.

关 键 词：创伤弧菌 溶细胞素 噬菌体展示 单链抗体 

分 类 号：R516[医药卫生—内科学]