H5N1亚型禽流感病毒血凝素蛋白单链抗体基因的克隆及表达  

Cloning and Expression of Single-Chain Fv Antibodies Against H5N1 Avian Influenza Virus Hemagglutinin

作　　者：郝淑美[1,2] 李群[1,2] 王承宇[2] 王化磊[2,3] 杨松涛[2] 夏咸柱[2] 

机构地区：[1]长春理工大学生命科学技术学院,长春130022 [2]解放军军事医学科学院军事兽医研究所,长春130062 [3]吉林大学畜牧兽医学院,长春130062

出　　处：《病毒学报》2009年第1期63-67,共5页Chinese Journal of Virology

基　　金：国家重点基础研究发展计划(973计划)课题(2005CB523005)

摘　　要：构建并表达H5N1亚型禽流感病毒血凝素蛋白单链抗体,为禽流感靶向治疗药物的研制制备靶向载体。从分泌血凝素单克隆抗体的杂交瘤细胞株中提取mRNA,采用RT-PCR法扩增出重链和轻链可变区基因,通过SOE-PCR法将重链和轻链通过Linker连接起来构建单链抗体基因,将获得的单链抗体基因装入原核表达载体pET28a(+)中,构建重组质粒并表达,以Western blot鉴定单链抗体的特异性。结果成功构建了单链抗体基因,全长714bp,经原核表达,所构建的单链抗体可与H5亚型禽流感病毒HA蛋白特异结合,为禽流感的靶向治疗奠定了基础。To construct Fv antibodies against H5N1 Avian influenza virus hemagglutinin,extracted mRNA from B lymphoblastoid cell lines secreting anti-HA antibodies was used and the VH and VL genes were amplified by RT-PCR and linked together by splicing overlap extension （SOE） with （Gly4 Ser）3 linker. The recombinant plasmid was then transformed to E. coli BL21 （DE3） and sequence analysis indicated the total length of scFv was 714bp and the expression of Fv was validated by PAGE and Western blot.

关 键 词：禽流感病毒 血凝素蛋白 单链抗体 克隆 表达 

分 类 号：S852.659[农业科学—基础兽医学]