Construction of recombinant replication competent plasmids containing HBV genome with mutations of precore and basic core promoter.
- Author:
Yan WANG
1
;
Dong JIANG
;
Lai WEI
;
Hong-song CHEN
;
Xu CONG
;
Ran FEI
;
Yu WANG
Author Information
- Publication Type:Journal Article
- MeSH: Carcinoma, Hepatocellular; metabolism; pathology; Cell Line, Tumor; Genetic Vectors; Genome, Viral; Hepatitis B Core Antigens; genetics; Hepatitis B virus; genetics; physiology; Humans; Liver Neoplasms; metabolism; pathology; Mutagenesis, Site-Directed; Plasmids; Recombinant Proteins; genetics; metabolism; Transfection; Viral Core Proteins; genetics; Virus Replication; genetics
- From: Chinese Journal of Experimental and Clinical Virology 2004;18(3):218-222
- CountryChina
- Language:Chinese
-
Abstract:
OBJECTIVETo construct a series of recombinant replication competent plasmids of hepatitis B virus (HBV) full-length genome with varying mutations of precore and basic core promoter.
METHODSThe plasmid containing the entire HBV (1.2 copies) genome was used to construct objective plasmids. The objective competent vectors were constructed by molecular cloning and PCR-based site-directed mutagenesis in vitro and confirmed with sequence analysis. After introducing the plasmids into hepatocellular carcinoma cell line (Huh7) by calcium phosphate transfection, the culture supernatant was collected to detect HBsAg and HBV DNA to analyze viral replication and expression.
RESULTSTen competent vectors with varying precore and basic core promoter mutations were constructed. After transfecting into Huh7, the vectors replicated and secreted viral particles.
CONCLUSIONSTen full-length competent HBV recombinants were obtained, which were harbouring varying mutations within precore and basic core promoter.
