ABSTRACT
Primers were synthesized according to DNA sequence of HBV found in China, and the reverse transcriptase (RT) region in polymerase gene, HBV preC/C region and the whole genome were amplified by PCR method from the serum of 8 patients with chronic HBV infection. Then the PCR products were ligated into pGEM Teasy vectors. 27 clones were sequenced. Deduced amino acid sequences were compared, and the results showed that mutations were widely distributed in structural and non structural viral proteins. The substitution mutations occur in clones isolated from specific patients, which led to a characteristic mutation in the quasispecies group. The results indicated that the diversity of HBV gene might play an important role in the pathogenesis of chronic HBV infection.
ABSTRACT
Polymerase chain reaction(PCR) was employed to amplify the whole HBV X region from the serum of patients with chronic hepatitis B virus(HBV) infection, and then the PCR products were subcloned into pGEM Teasy vectors. Clones were randomly selected to be sequenced. Comparison of the cloned sequence was made to look for the differene. The sequencing results showed that each sequence of selected clones was differenct and there were HBV quasispecies groups in patients. There was hot deletion region near the 3' end of X gene. To address whether the mutations were responsible for the transactivating effect,the wild type(wt) and the mutants of HBV X genes were subcloned into the EcoRI site of pcDNA3 1( ) vectors, and the recombinant plasmids were cotransfected into HepG2 cells with reporter plasmid pSV lacZ,respectively. The activity of ? galactosidase controlled by SV40 early promoter/enhancer was detected, which reflected the transactivating function of HBx protein. The cotransfection results indicated that the wt HBV X gene acted as a transactivator on the SV40 early promoter/enhancer, and the mutations which caused premature termination of the X open reading frame reduced their transactivating effects to certain extent.
ABSTRACT
To investigate the HBV quasispecies groups in the patients with chronic HBV infection. A set of specific primers was synthesized according to HBV DNA sequence of the Chinese strain, and the whole pre core/core region was amplified by PCR method from the serum of 4 patients with chronic HBV infection.Then the PCR products were subcloned into pGEM Teasy vectors. Clones were selected to be sequenced,and then sequence comparison was made to find the difference. Each sequence of selected clones was found to be different. The homology of the nucleic acid sequence from the same patient was above 98 0%. There were many mutation types in this region, including point substitution, core region internal deletion, as well as frame shift reading. The results indicated that there were HBV quasispecies groups in chronically infected patients. The mutation in this region may influence the prognosis of HBV infection.