Complete HBV DNA clone and sequence from serum samples of severe hepatitis B patients.
- Author:
Wei WU
1
;
Lan-juan LI
;
Yu CHEN
;
Jun LI
;
Xiang QIAN
;
Dong-qing CHENG
Author Information
- Publication Type:Journal Article
- MeSH: Adult; B-Lymphocytes; metabolism; Cloning, Molecular; Epitopes; genetics; Genome, Viral; genetics; Hepatitis B; etiology; virology; Hepatitis B Surface Antigens; genetics; Hepatitis B virus; genetics; Humans; Male; Middle Aged; Mutation; Protein Precursors; genetics; Sequence Analysis; T-Lymphocytes, Cytotoxic; metabolism
- From: Chinese Journal of Hepatology 2005;13(10):734-737
- CountryChina
- Language:Chinese
-
Abstract:
OBJECTIVETo study the association between hepatitis B virus (HBV) mutants and the pathogenesis of severe hepatitis B by full-length HBV genome.
METHODSSerum 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.
RESULTS4 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.
CONCLUSIONSThis method could serve to study the relationship between HBV genome and the pathogenesis of severe hepatitis B.
