FMDV整联蛋白受体β1亚基的克隆及其配体黏附区多抗的制备  

Cloning of Foot-and-Mouth Disease Virus Integrin Receptor β_1 Subunit and Antibody Production to Its Ligand-binding Domain

作　　者：杜平[1] 尚佑军[1] 马军武[1] 贺延玉[2] 孙晓林[3] 刘湘涛[1] 

机构地区：[1]中国农业科学院兰州兽医研究所家畜疫病病原生物学国家重点实验室农业部畜禽病毒学重点实验室,兰州730046 [2]甘肃农业大学研究测试中心,兰州730070 [3]甘肃农业大学动物医学院,兰州730070

出　　处：《生物工程学报》2008年第5期874-880,共7页Chinese Journal of Biotechnology

基　　金：国家科技支撑计划(No.2006BAD06A03)~~

摘　　要：采用RT-PCR技术从牛气管组织扩增出2400bp的β1基因,回收纯化连入PGEM-T载体,测序。用Expasy软件对β1基因的抗原性进行分析,选取胞外区334-861bp的配体结合区与6×His融合,在大肠杆菌中大规模诱导表达,并经Ni2＋亲和柱层析纯化。通过SDS-PAGE鉴定后,应用纯化蛋白免疫新西兰家兔,获得效价在1：12800以上的多抗,Western blotting鉴定表明此抗体可特异性的与表达的融合蛋白作用。We produced β1 gene which is about 2400 bp by reverse transcription polymerase chain reaction （RT-PCR） from bovine trachea, reclaimed and purified, then cloned the amplified fragment to pGEM-T easy vector, confirmed by sequencing. The immuno-dominant epitope of β1 gene was chosen by computer analysis and then syncretized ligand-binding domain from 346 bp to 843 bp of ecytoplasm with six histidine, expressed LBD protein massly in E. coli BL21 （DE3）, and identified by SDS-PAGE. The fusion protein was purified with Ni-NTA affinity chromatography and immunized New Zealand rabbits preparing of its polyclonai antibody, the specific antibody titer was above 1：12 800 detected by indirect ELISA, the result of Western blot showed that this antibody could be recognized by LBD fusion protein.

关 键 词：口蹄疫病毒 整联蛋白 Β1亚基 配体结合区 多克隆抗体 

分 类 号：S852.65[农业科学—基础兽医学]