用于大容量噬菌体抗体库的表达载体的构建及鉴定  

Construction of a vector suitable for making large phage antibody library

作　　者：王琰[1] 王欲晓[1] 陈晓穗[1] 化冰[1] 刘晓琳[1] 

机构地区：[1]解放军海军总医院,北京100037

出　　处：《中国免疫学杂志》2003年第2期93-96,共4页Chinese Journal of Immunology

基　　金：解放军医药卫生"十五"重点项目(012015)资助课题

摘　　要：目的:构建一个适用于大容量抗体库的噬菌体抗体表达载体。方法:通过插入人工合成寡核苷酸、PCR介导的定位突变等技术,将载体p3MH进行改造,使之更适于构建大容量抗体库。通过噬菌体抗体的表达、ELISA及细胞内重组试验鉴定所获得的表达载体。结果:通过插入loxp511和loxp序列,将Lac启动子更换为阿拉伯糖启动子、改造抗体基因克隆位点等,将p3MH改造成pAL。经检测pAL可以表达具有功能的Fab噬菌体抗体,其表达调控更为严密,可以在cre+细菌胞内发生预期的loxp-cre介导的定位重组。结论:所获得的表达载体pAL适用于构建大容量Fab噬菌体抗体库。Objective:To construct a vector that suits the construction of large phage antibody libraries.Methods:Hie phage antibody vector p3MH was modified by insertion of synthesized oligos and PCR mediated site-specific mutations. Hie resultant vector was checked by expression of phage antibody, ELBA and in vivo recombination. Results: Vector pAL was obtained by following modifications of p3MH: insertion of Ioxp511 and loxp sequences, substitution of Lac promoter by Ara promoter, and modification of the cloning site for antibody genes. pAL was proved capable of expressing functional Fab phage antibodies under tighter control. In ere+ bacteria, pAL exhibited loxp-cre mediated recombination. Conclusion: pAL is useful for construction of large phage antibody libraries.

关 键 词：噬茵体抗体库 容量 表达载体 p3MH PAL loxp-cre定位重组系统 

分 类 号：Q78[生物学—分子生物学]