Identification of a novel negative regulatory element on the hepatitis B virus S-(+)-strand

In this study, we scanned the whole hepatitis B virus (HBV) genome for the identification of potential regulatory elements located on the S-(+)-strand. With pCDNA3.1-HBV1.3 as template which contains 1.3-fold HBV whole genome, HBV fragments were amplified by PCR methods, and then inserted into the upstream of a heterologous luciferase reporter vector (pGL3control) in antisense orientation, allowing the HBV expression from the S-(+)-strand. We found that the reporter plasmid containing nt 509-1(3182)-2639 of HBV inhibited luciferase gene transcription and expression in HepG2 cells. Our results strongly suggested that nt 453-250 of HBV may act as a novel negative regulatory element, which has not been reported before. Serial deletion analyses further indicated that nt 453-250 sequence of HBV genome would be the minimal sequence essential for the inhibitory effect of the novel negative regulatory element.