Genomics and Applied Biology 2016, Vol.7, No.5, 1-8
1
Research Article Open Access
Research on HBV Gene Integration into Host Genome that is Related to HBV
DR Region
Haoxiang Luo
, Long Gu, Lihong He
Department of medicine, Southwest of Guizhou Vocational and Technical College for Nationalities, Xingyi 562400, China
Corresponding author Email
Genomics and Applied Biology, 2016, Vol. 7, No.5 doi
Received: 4 Jul., 2016
Accepted: 6 Sep., 2016
Published: 9 Nov., 2016
Copyright © 2016
Luo et al., This is an open access article published under the terms of the Creative Commons Attribution License, which permits
unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.
Preferred citation for this article
:
Luo H.X., Gu L., He L.H., 2016, Research on HBV Gene Integration into Host Genome that is related to HBV DR region, Genomics and Applied Biology, 7(5):
1-8 (doi
Abstract
In most studies, the HBV DNA has been found integrated into the DNA of the hepatocellular carcinoma cells, the integrated HBV
DNA is related to HBV DR region, in order to research how HBV integrated into the genome of the cells via HBV DR region, we
isolated HBV Creg DNA fragment which is from nt1087 to nt2488 contained the DR region, regulatory sequence, X gene and C gene
and constructed pcDNA3.1(+)-HBV Creg eukarya expression vector, then transfected it into the HepG2 cells to observe the
integration way of HBV. In this study, HBV Creg DNA fragment was isolated from HBV genome by recombinant PCR, in this DNA
fragment, the DR region, regulatory sequence, X gene and C gene of HBV was included, then linked it to the vector pcDNA3.1(+) to
construct the pcDNA3.1(+)-HBV Creg eukarya expression vector. We transfected the pcDNA3.1(+)-HBV Creg eukarya expression
vector into HepG2 cells, in the process, the transfected cell lines were selected by G418, the total DNA of HepG2 cells was extracted
to test the integration of HBV by PCR using different primers. The full length of HBV Creg DNA fragment contained DR region
could only be detected by PCR before integration into cell genome, after the fragment integrated into HepG2 genome, the full length
of HBV Creg DNA fragment had not been detected all the time, however, HBV X gene and HBV C gene could be detected
respectively, for the reason of HBV DR region, it may integrate at DR1 or DR2 with the Creg DNA extending downstream or
upstream though C gene or X gene. The HBV Creg fragment can integrate into HepG2 cells genome. And the integration of HBV is
related to DR region, the way of HBV Creg DNA fragment integrated into HepG2 cells is from DR region to both sides.
Keywords
Hepatitis B Virus; Hepatocellular carcinoma; Gene Recombinant; DR Region; Gene integration
1 Introduction
In most cases of HCC (HCC, hepatocellular carcinoma), the HBV DNA has been found integrated into the DNA
of the tumorous hepatocytes (Di Bisceglie, 2009). And the integrated HBV DNA can exist as whole genomes or
reorganized subgenomic fragments. Dejean et al. sequenced the host-viral DNA junctions of two integrated HBV
DNA sequences cloned from two different human liver tumors (Dejean et al., 1984). In each clone, one junction
mapped to within one of two identical 11- base-pair (bp) 5' T-T-C-A-C-C-T-C-T-G-C cohesive - end sequences
(DR1 and DR2) that are directly repeated near the extremities of the viral (+) and (-) strands, which start at
nucleotides 1590 and 1824, were termed DR (DR, direct repeat region)1 and DR2, respectively. It is known by us
that this HBV DR region was a crucial region about the integration of HBV, but it is also a crucial region about the
replication of HBV. Between DR1 and DR2 region, there exists a gap, when the virus replicates, the gap will be
filled up ahead of time. As far as the integration site of the viral DNA is concerned, reconmbinational mechanism
via the single - stranded gap region of the viral genome have been proposed the integrated HBV DNA appears to
have one fixed end at the virus - cell junction within the DRs. However, the function of this region was known
very little by us, and the formation mechanism of HBV cccDNA (covalently closed circular DNA) and the
integration mechanism of HBV DR region has not yet been reported, so the research on this region is very
important. So we constructed the pcDNA3.1(+) - HBV Creg eukarya expression vector, in this Creg fragment, it
contained DR region, X gene, C gene, and the regulatory sequence, then we made it transfect into HepG2 cells to
find out the integration mechanism of HBV DR region.
