Expression and antigenic analysis of three recombinant continuous ORF2 proteins of swine hepatitis E virus Guangxi strain

In order to determine the distribution of hepatitis E virus(HEV) ORF2 antigenic fragments,two pairs of primers were designed according to the sequence of China T1 strain and a fragment of ORF2 gene was amplified from fresh porcine feces.The fragment named as HEVORF2 A was 1 725 bp in size,located between nucleotide 205 and 1 929 of ORF2.Subsequently,three pairs of primers were designed based on HEVORF2 A,and three continuous gene fragments were amplified from HEVORF2-A,which were named as HEVORF2-B1,HEVORF2-B2 and HEVORF2-B3,and corresponded with the nucleotide 1 to 576,577 to 1149 and 1150 to 1725 in HEVORF2-A,respectively.The above four gene fragments were cloned into the expression vector pET32a(+),and the recombinant expression plasmids were transformed into competent cells BL21(DE3) pLysS and induced with IPTG,respectively.Three fusion proteins of 41.3 ku,41.5 ku and 41.8 ku in molecular weight were highly expressed from pET32a-B1,pET32a-B2 and pET32a-B3,respectively,but only a few of protein of 82.6 ku was expressed from pET32a-A.Western-blot analysis showed that the 3 short fusion proteins could be recognized by the sera from the naturally-infected pigs and the positive control sera from the HEV ELISA Kit of Beijing Wantai Coporation,indicating that the 3 short fusion proteins expressed from the 3 constructed expression vectors had good antigenicity.