甲型H1N1流感病毒血凝素单克隆抗体轻链和重链可变区基因的巢式PCR扩增及序列分析  

Nested PCR Amplifying and DNA Sequence Analysis of Light and Heavy Chain Variable Region Gene of Monoclonal Antibody Against H1N1 Influenza Virus Hemagglutinin

作　　者：李慧瑾[1] 李研[1] 孙晶莹[1] 高锦伟[1] 赵向绒[1] 封青[1] 谭天天[1] 胡巧侠[1] 李元[1] 胡军[1] 

机构地区：[1]陕西省人民医院中心实验室,陕西西安710068

出　　处：《生物技术通讯》2013年第1期61-64,共4页Letters in Biotechnology

基　　金：国家自然科学基金(81202373)

摘　　要：目的:采用巢式PCR对甲型H1N1流感病毒血凝素单克隆抗体的轻链和重链基因进行扩增,对获得的基因进行序列分析,并找出克隆鼠Igκ轻链和重链可变区基因的通用方法。方法:设计22对扩增鼠Igκ轻链可变区和重链可变区基因的引物,对6株鼠抗人甲型H1N1流感病毒血凝素单克隆抗体的轻链和重链可变区基因进行克隆并测序,与NCBI公布的鼠免疫球蛋白序列比对分析。结果:巢式PCR方法可以有效避免单克隆抗体克隆过程的假基因,并且得到的单克隆抗体的氨基酸序列均符合鼠免疫球蛋白可变区特征。结论:建立了克隆鼠免疫球蛋白轻链和重链可变区基因的通用方法,为后期克隆鼠源性单克隆抗体的可变区基因提供了基础,并为研究甲型H1N1流感病毒血凝素与抗体的结合位点提供了实验数据。Objective： To clone and analysis light and heavy chain genes of monoclonal antibody against H1N1 influenza virus hemagglutinin by Nested PCR, then construct a common method for cloning light and heavy chain variable region genes of mouse anti-human H1N1 influenza virus hemagglutinin monoclonal antibody. Methods： We designed 22 pairs of primers for amplifying IgK light and heavy chain variable region gene, and cloned light and heavy chain variable region genes of 6 mouse anti-human H1N1 influenza virus hemagglutinin monoelonal anti- bodies. Cloning and subsequent sequences analysis of immunoglobulin gene were performed, sequence alignment with mouse immunoglobulin in NCBI. Results： The nucleotide and corresponding amino acid sequences were in line with characteristics of mouse immunoglobulin variable region. Conclusion： In this study, we cloned 6 mouse antibody variable region genes, and found a common method for cloning immunoglobulin light and heavy chain variable region genes. It provides a basis for late amplifying monoelonal antibody variable region, and experimental data for analysis of H1N1 influenza virus hemagglutinin and antibody binding sites.

关 键 词：甲型H1N1流感病毒 单克隆抗体 可变区 巢式PCR 序列分析 

分 类 号：Q78[生物学—分子生物学] R392.1[医药卫生—免疫学]