食品科学2011，Vol.32Issue(15)：200-204,5.
抗克伦特罗核糖体展示单链抗体文库的构建及鉴定
Construction and Identification of an Anti-clenbuterol Single-chain Fv Library for Ribosome Display
摘要
Abstract
In order to construct a high-capacity ribosome display anti-clenbuterol single chain Fv（CBLscFv） library for the selection of high affinity scFv antibody,CBL-BSA immunogen was immunized to Balb/c mice.Spleen cells were isolated and total RNA was extracted by the Trizol method for cloning heavy chain and light chain gene by RT-PCR.VH and VL were rearranged randomly by SOEing（splicing by overlap extension）.Finally,the elements T7 promoter,3＇ stem loop structure,ribosome binding sites,spacers and 5＇ stem loop structure were screened in vitro.The constructed library was verified by blue/white screening and further sequencing.The results showed that the titer of antiserum from immunized mice for library construction was 1：128000.The heavy and light chain variable region genes were successfully amplified to be approximately 360 bp and 330 bp,respectively.The fragment of the constructed ribosome display library was approximately 1200 bp with a capacity of 1.75 × 1013.Meanwhile,sequence analysis was carried out using IMGT/V-QUEST database.The results showed that the heavy chain and light chain variable region genes were rearranged by mouse IG set,and the homology rates were more than 84%.Amino acid sequence alignment revealed a homology rate of 37.82% for heavy chain amino acid sequences and 39.09% for light chain amino acid sequences.The diversity was observed in VH-CDR3,VL-CDR1,VL-CDR2 and VL-CDR3.Therefore,a high-capacity ribosome display library with the function for antibody screening was successfully constructed.关键词
核糖体展示/克伦特罗/单链抗体文库Key words
ribosome display/clenbuterol/single-chain fragment variant（scFv）分类
生物科学引用本文复制引用
刘细霞,孙远明,董洁娴,杨武英,谢曦,王弘..抗克伦特罗核糖体展示单链抗体文库的构建及鉴定[J].食品科学,2011,32(15):200-204,5.基金项目
国家自然科学基金项目（30871755 （）
20877029） （）
