Construction of a replication-competent hepatitis B virus vector carrying secreted luciferase transgene and establishment of new hepatitis B virus replication and expression cell lines

Figure 3 Establishment of secretory Nanoluc Luciferase reporter gene cell lines carrying stably secreted hepatitis B virus particles. HepG2 TA-7 cells were transfected with the replication-competent vector pTRE-sNLuc, and the isolated clone of cells was selected in the presence of hygromycin. A: Selection of cell lines with good luciferase activity. After several passages, the Nos. 1, 7, 22, 23, and 35 cell lines were screened, and luciferase expression in the supernatant was detected with Nano-Glo luciferase assay reagent; B: Selection of cell lines with good hepatitis B virus (HBV) replication. Extracellular viral particles of cell lines Nos. 7, 23, and 35 were analyzed by Southern blot using a ³²P-labeled HBV probe. The relaxed circular DNA and double-stranded linear DNA signals are indicated; C: Infection of HepaRG cells by secretory Nanoluc Luciferase recombinant HBV particles. Differentiated HepaRG cells were inoculated with viral particles from supernatants of cell lines No. 35 or not (negative control), and luciferase expression in the supernatant was detected at the indicated time points (0, 2, 4, 6, 8, and 10 d). HBV: Hepatitis B virus; RLU: Relative light unit.