禽流感病毒PA基因的原核表达载体的构建及鉴定  

Construction of prokaryotic expression vector of PA gene of avian influenza virus and identification of the expression vector

作　　者：赵翠君[1] 黄会岭[1] 杜聚朝 王连群 李云 高慧芳 

机构地区：[1]河北农业大学动物科技学院,河北保定071001 [2]邯郸市动物卫生监督所,河北邯郸056001 [3]石家庄市畜产品质量监测中心,河北石家庄050011 [4]河北省迁安市农业中心,河北迁安064400

出　　处：《河北农业大学学报》2008年第6期95-98,103,共5页Journal of Hebei Agricultural University

摘　　要：以重组质粒pcDNA-PA为模板扩增PA全基因,并构建原核表达载体pGEX-6p-1-PA。根据GenBank公开发表的禽流感病毒PA基因的序列设计引物,用PCR方法从重组质粒pcDNA-PA中扩增禽流感病毒的PA基因,将该基因定向插入到原核表达载体pGEX-6p-1中,然后将连接产物转化宿主菌DH5α,挑取阳性克隆进行PCR和BamHⅠ、NotⅠ双酶切鉴定并测序。测序结果表明,禽流感病毒PA基因全长2151 bp,编码717个氨基酸,原核表达载体pGEX-6p-1-PA已构建成功。从而为禽流感蛋白PA的表达及其多克隆抗体的制备奠定了基础。The complete PA gene of Avian Influenza Virus （AIV） was amplified with recombinant plasmid pcDNA- PA as template, and prokaryotic expression vector pGEX - 6p - 1 - PA was con- structed. According to the relevant nucleotide sequence from GenBank, a pair of specific primers were designed, and the complete PA gene of AIV was amplified with recombinant plasmid pcDNA - PA as template by PCR method. The PCR fragment was inserted into pGEX - 6p - 1. The ligat- ed product was transformed into E. coli DHSa competent cells. The positive clones were identified by PCR, digestion with BarnH I and Not I, and sequence. Sequence analysis demonstrated that PA gene contained one open reading frame of 2 151 bp, which encoded a polypeptide of 717 amino acids, therefore the prokaryotic expression vector pGEX - 6p - 1 - PA was constructed successfully. This established a foundation for expression of AIV PA protein and preparation of its polyclonal antibody.

关 键 词：禽流感病毒 PA基因 原核表达载体 

分 类 号：S852.65[农业科学—基础兽医学]