乙型肝炎病毒全基因重组真核表达质粒的构建

Construction of recombinant eukaryotic expression plasmid containing full-length genome of hepatitis B virus

目的 构建乙型肝炎病毒（ＨＢＶ）全基因重组真核表达质粒，旨在转染肝细胞，建立 ＨＢＶ复制状态模型。方法体 外连接 ＨＢＶ（ａｙｗ亚型） ＤＮＡ获得 ６．４ ｋｂ的双拷贝 ＤＮＡ片段，然后将其插入 ｐｃＤＮＡ３载体的 ＥｃｏＲＶ位点。结果通过 聚合酶链反应（ＰＣＲ）和酶切筛选和验证获得头尾串联双拷贝ＨＢＶ全基因重组真核表达质粒。结论成功构建了ＨＢＶ 全基因重组真核表达质粒。

Objective To construct an recombinant eukaryotic expression plasmid containing tandem copies of full-length hepatitis B virus (HBV) genome, to establish models simulating HBV replication status in the liver cells. Methods The DNA fragment of the 2 copies of HBV (ayw subtype) genome was obtained in vitro and inserted into EcoRV site of the plasmid pcDNA3. Results After screening with PCR and restriction enzyme analysis, the recombinant eukaryotic expression plasmid containing tandem copies of HBV genome in head-to-tail connection was obtained. Conclusion The recombinant eukaryotic expression plasmid is successfully constructed.