Published online Dec 21, 2015. doi: 10.3748/wjg.v21.i47.13360
Peer-review started: June 11, 2015
First decision: July 10, 2015
Revised: July 17, 2015
Accepted: September 30, 2015
Article in press: September 30, 2015
Published online: December 21, 2015
Processing time: 190 Days and 0.5 Hours
AIM: To develop a Fok-I nested polymerase chain reaction (PCR)-restriction fragment length polymorphism analysis (PRA) method for the detection of hepatitis B virus X region (HBx) V5M mutation.
METHODS: Nested PCR was applied into DNAs from 198 chronic patients at 2 different stages [121 patients with hepatocellular carcinoma (HCC) and 77 carrier patients]. To identify V5M mutants, digestion of nested PCR amplicons by the restriction enzyme Fok-I (GGA TGN9↓) was done. For size comparison, the enzyme-treated products were analyzed by electrophoresis on 2.5% agarose gels, stained with ethidium bromide, and visualized on a UV transilluminator.
RESULTS: The assay enabled the identification of 69 patients (sensitivity of 34.8%; 46 HCC patients and 23 carrier patients). Our data also showed that V5M prevalence in HCC patients was significantly higher than in carrier patients (47.8%, 22/46 patients vs 0%, 0/23 patients, P < 0.001), suggesting that HBxAg V5M mutation may play a pivotal role in HCC generation in chronic patients with genotype C infections.
CONCLUSION: The Fok-I nested PRA developed in this study is a reliable and cost-effective method to detect HBxAg V5M mutation in chronic patients with genotype C2 infection.
Core tip: In the present study, we developed a reliable and cost-effective Fok-I nested polymerase chain reaction-restriction fragment length polymorphism analysis (PRA) method for the detection of V5M from chronic patients with genotype C2 infection. In addition, our epidemiological data based on the Fok-I nested PRA method strongly support the previous reports that V5M may play a very pivotal role in hepatocarcinogenesis, at least in chronic patients infected with genotype C2.