Molecular cloning of HBV S-protein mutants for expression in human HepG2 liver cell line

Abstract

Occult hepatitis B virus (HBV) infection (OBI) is defined as low plasma level of HBV DNA with undetectable HBV surface antigen (HBsAg). Multiple amino acid substitutions in the S protein may affect the HbsAg secretion and detection by commercial assays. In the current study, a cell culture system was optimized and will be used to study the potential role of OBI specific amino acid substitutions in the S protein. A total of 10 HBV mutants were selected from OBI blood donors. Surface gene amplification using gene specific primers were cloned in a plasmid, transformed in competent cell line and transfected in HepG2 cells. After several attempts of optimization, successful cloning method and culturing of HepG2 cell line were achieved. Optimized protocols for transfecting mutant HBsAg are currently available and will be used to study the in vitro effect of HBV amino acid substitutions.