皮蝇Hypodermin A基因的克隆及表达质粒构建  

Cloning and Construction of Recombinant Expression of Hypodermin A Tang

作　　者：唐立刚[1] 杨东英[1] 罗建勋[2] 欧阳五庆[1] 殷宏[2] 

机构地区：[1]西北农林科技大学动物科技学院,陕西杨陵712100 [2]中国农业科学院兰州兽医研究所,甘肃兰州730046

出　　处：《动物科学与动物医学》2004年第1期22-23,19,共3页Animal Science & Veterinary Medicine

基　　金：国家863项目(2001AA249081);欧盟资助项目(ICA4-CT-2000-30028)

摘　　要：参照牛皮蝇HypoderminA(HA)基因的核苷酸序列,设计一对引物,以皮蝇总RNA为模板进行RT--PCR扩增牛皮蝇HA基因,将扩增基因进行克隆测序。序列分析表明,所克隆获得的基因与GenBank中已经登录的核苷酸和氨基酸的同源性分别为97.2%和99.3%。同时,将该基因与表达载体PGEX-4T-1连接,构建并获得阳性重组表达载体。Total RNA was extracted from Hypoderma,then mRNA was isolated by using oligo(dT)as a probe.The HA gene specific primers were devised by GOLDkey software.A700bp specific fragment was amplified by RT-PCR and ligated into PGEM-T Easy vector.It was identified by restriction endonuclease analysis and PCR and sequencing that this fragment contained the complete open reading frame (ORF)of the HA gene.In comparison with GenBank data,the homologies of the nucleotide sequence and amino acid sequence are97.2%and99.3%,respectively.After the HA-T was restrictively digested,the interested gene HA was subcoloned into the prokaryotic expression vector PGEM-4T-1.Positive clones were selected by restriction endonuclease analysis and PCR identification.

关 键 词：牛皮蝇 HYPODERMIN A基因 基因克隆 基因表达 皮蝇蛆病 基因工程疫苗 

分 类 号：S852.7[农业科学—基础兽医学]