摘要
目的构建戊型肝炎病毒(HEV)ORF3真核表达载体pcHEV3并进行初步鉴定。方法从实验感染HEV新疆株的猕猴胆汁中提取HEV RNA,逆转录法合成HEV cDNA,采用RT-PCR方法扩增HEV ORF3 cDNA片段,将其克隆至载体质粒pcDNA3,构建HEV ORF3真核表达载体,经抗生素初步筛选后,重组阳性载体进行酶切和测序鉴定。结果从实验感染HEV新疆株的猕猴胆汁中克隆出HEV ORF3全长cDNA片段,经酶切鉴定及DNA测序鉴定,成功构建HEV ORF3蛋白真核表达载体pcHEV3。结论成功构建HEV ORF3蛋白真核表达载体,为进一步研究HEV ORF3蛋白的功能提供了条件。
Objective To construct the eukaryotic expression vector carrying hepatitis E virus （HEV） ORF3 cDNA and to identify preliminarily.Methods HEV RNA was extracted from the bile of macaque experimental infected with HEV.Then HEV cDNA was synthesized through reverse-transcription reaction with HEV RNA as template.HEV ORF3 cDNA was amplified by PCR and cloned into plasmid pcDNA3.HEV ORV eukaryotic expression vector was constructed,then we used antibiotics for preliminary screening.The positive recombinant vector was identified by enzyme digestion and sequencing.Resuits After the enzyme digestion and sequencing,the HEV ORF3 protein eukaryotic expression recombinant vector pcHEV3 was successfully constructed.Conclusions The HEV ORF3 protein eukaryotic expression recombinant vector is successfully constructed,which provides experimental foundation for exploring the biological function of HEV ORF3 protein.
出处
《山东医药》
CAS
2014年第2期17-19,共3页
Shandong Medical Journal
基金
河南省教育厅自然科学研究计划项目(2010C310005)
