Complete HBV DNA clone and sequence from serum samples of severe hepatitis B patients / 中华肝脏病杂志

ABSTRACT

<p><b>OBJECTIVE</b>To study the association between hepatitis B virus (HBV) mutants and the pathogenesis of severe hepatitis B by full-length HBV genome.</p><p><b>METHODS</b>Serum samples from 10 severe hepatitis B patients were collected in our hospital. Serum HBV DNAs were extracted using DNA mini Kit, and amplified by LA Taq DNA polymerase to yield full-length HBV DNA. PCR products were isolated and cloned into vector pUCm-T, then transfected into DH-5 alpha cells. Positive clones were selected and checked by digestion, and full-length HBV DNAs were sequenced.</p><p><b>RESULTS</b>4 cases were cloned into vector pUCm-T successfully and completed the full-length sequencing. Among them, 3 cases had a G to A mutation at nucleotide 1896 in pre-C region and 1 had a double mutation of T1762-A1764 in the core promoter region. Some amino acid changes occurred within the known CTL, B or T cell epitopes of the PrS2 and C regions.</p><p><b>CONCLUSIONS</b>This method could serve to study the relationship between HBV genome and the pathogenesis of severe hepatitis B.</p>