[Effects of different vectors and gene fragments on antigen expression of hepatitis E virus DNA immunization].
OBJECTIVE
To investigate the effects of different vectors and gene fragments on antigen expression of hepatitis E virus (HEV) DNA immunization.


METHODS
Gene fragments encoding p166 and p179, which contain the neutralization antigenic epitopes of a Chinese strain of HEV genotype IV, were cloned into two different eukaryotic expression vectors (pTR421 and pCDNA3.1), respectively. The in vitro expression level of p166 and p179 in HepG2 cells transfected by each of the recombinant plasmids with lipofectamine2000 was examined by means of immunofluorescence and Western blot. Meanwhile, the in vivo expression level in muscles of mice was examined with immunohistochemistry staining.


RESULTS
Four recombinant plasmids, pTR421-166, pTR421-179, pCDNA3.1-166 and pCDNA3.1-179, were constructed successfully and confirmed correct with restriction endonuclease analysis and nucleotide sequencing. The antigen expression was only detected in HepG2 cells transfected by pTR421-179 and in myocytes of the mice injected with pTR421-179. Neither in vitro nor in vivo antigen expression was detected with pTR421-166 although p166 was only 13 amino acids shorter than p179 at N terminus. Neither pCDNA3.1-166 nor pCDNA3.1-179 was expressed in vitro and in vivo.


CONCLUSIONS
Selection of the vectors and gene fragments is critical to HEV gene expression and HEV DNA vaccine.
Fang
Xue-feng
XF
Department of Pathogen Biology and Immunology, School of Basic Medical Sciences, Southeast University, Nanjing 210009, China. fxfy2000@sohu.com
Wang
Li-xin
LX
Meng
Ji-hong
JH
Dong
Chen
C
Tian
Hua
H
Zhai
Li-jie
LJ
D000818:Animals; D015153:Blotting, Western; D004279:DNA, Viral / Q000235:genetics* / Q000276:immunology*; D005260:Female; D005455:Fluorescent Antibody Technique; D005786:Gene Expression Regulation; D005822:Genetic Vectors / Q000235:genetics*; D016752:Hepatitis E virus / Q000235:genetics* / Q000276:immunology* / Q000378:metabolism; D051379:Mice; D008807:Mice, Inbred BALB C; D016133:Polymerase Chain Reaction; D019444:Vaccines, DNA / Q000235:genetics*
D004279:DNA, Viral; D019444:Vaccines, DNA