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Stable expression of HBV C gene mutants in immortalized human B-cell lines / 南方医科大学学报
Article in Chinese | WPRIM | ID: wpr-282932
Responsible library: WPRO
ABSTRACT
<p><b>OBJECTIVE</b>To provide an cell model of immortalized lymphoblstoid B-cell lines for studying the biological characteristics of full-length hepatitis B virus (HBV) genome carrying the hot-spot mutations V60, G87, and L97.</p><p><b>METHODS</b>V60, G87, and L97 mutation points were introduced into HBV p3.8 II plasmid containing 1.2 copy of HBV genome by means of site-directed mutagenesis. The HBV genome was amplified by PCR from p3.8 II and p3.8 II-V60, G87, L97 plasmid, and the PCR product was inserted into EBO-plpp eukaryotic expression vector. The recombinant vectors and the EBO-plpp vector were transfected into immortalized human lymphoblasts with lipofectamine 2000 and selected with hygromycin. Steady expression of the target genes was determined by RT-PCR, Western blotting and microparticle enzyme immunoassay.</p><p><b>RESULTS</b>DNA sequence analysis indicated that the desired mutation was introduced into wild-type HBV DNA. HBsAg, HBeAg and HBcAg could be detected in EBO-HBV-transfected cell lysate or culture supernatant.</p><p><b>CONCLUSION</b>Transfectants that stably express HBV mutant antigen may provide a cell model to study the biological characteristics of HBV carrying hot-spot mutation in vitro.</p>
Subject(s)
Full text: Available Index: WPRIM (Western Pacific) Main subject: Virology / Humans / DNA, Viral / Cell Line, Transformed / Molecular Sequence Data / B-Lymphocytes / Base Sequence / Transfection / Gene Expression Regulation, Viral / Cell Transformation, Viral Language: Chinese Journal: Journal of Southern Medical University Year: 2006 Type: Article
Full text: Available Index: WPRIM (Western Pacific) Main subject: Virology / Humans / DNA, Viral / Cell Line, Transformed / Molecular Sequence Data / B-Lymphocytes / Base Sequence / Transfection / Gene Expression Regulation, Viral / Cell Transformation, Viral Language: Chinese Journal: Journal of Southern Medical University Year: 2006 Type: Article