Cloning and Prokaryotic Expression of cDNAs from Hepatitis E Virus Structural Gene of the SW189 Strain

It is necessary to study the second open reading frame (open reading frame 2, ORF2) and antigenic peptidescharacteristic of genotype 4 of HEV, and create one kind of high sensitivity and specificity of anti-HEV IgM detectionkit. The objective of this study was to obtain recombinant antigen for development of anti-HEV ELISA method andvaccine against hepatitis E virus infection. A 728 base cDNA was collected from 5'-terminus of open reading frame 2(ORF2) among epidemic hepatitis E virus (HEV) isolated from Gansu, Western China. The fragment was digestedwith Not I and Nco I, and inserted into vector pET32a(+). The recombinant plasmid was transformed into E. coliRosetta and the fusion protein expressed was confirmed by Western blot analysis. The recombinant plasmid wasidentified and confirmed with enzyme digestion, polymerase chain reaction (PCR) and sequencing, respectively. Aprotein band of about 45.3 kDa was demonstrated by SDS-PAGE. The result of Western blot analysis suggested thatthe fusion protein reacted with anti-HEV positive sera at a dilution of 1:1500. The recombinant protein pORF2 may beuseful in developing anti-HEV ELISA kit and vaccine against hepatitis E virus infection.