Cloning and expression of cDNAs from hepatitis E virus structural gene.
OBJECTIVE
To obtain recombinant antigen for development of anti-HEV ELISA kit and vaccine against hepatitis E virus infection.


METHODS
A 492 base cDNA was collected from 5-terminus of open reading frame 2 (ORF2) among epidemic hepatitis E virus (HEV) isolated from Xinjiang, Western China. The fragment was digested with BamH I and EcoR I, and inserted into vector pGEX-4T-3 which was also digested by the same enzyme. The recombinant plasmid was transformed into E.coli TG-1 and the fusion protein expressed was confirmed by Western blot analysis using serum of a patient with hepatitis E.


RESULTS
The recombinant plasmid was identified and confirmed with enzyme digestion, polymerase chain reaction (PCR) and sequencing, respectively. A protein band of about 46 kDa was demonstrated by SDS-PAGE and designated GST-pORF2. The result of Western blot analysis suggested that the fusion protein reacted with anti-HEV positive sera at a dilution of 1:100.


CONCLUSIONS
The recombinant protein GST-pORF2 may be useful in developing anti-HEV ELISA kit and vaccine against hepatitis E virus infection.
Ruan
Bing
B
First Affiliated Hospital, Medical College of Zhejiang University, Hangzhou 310003, China. my_bing@mail.hz.zj.cn
Zumbika
Edward
E
Wang
Shu-Ying
SY
Chen
Yong
Y
Ma
Yi-Lin
YL
Chen
Ya-Gang
YG
Liu
Ke-Zhou
KZ
D000818:Animals; D003001:Cloning, Molecular; D018076:DNA, Complementary; D004926:Escherichia coli; D015967:Gene Expression Regulation, Viral; D016752:Hepatitis E virus / Q000235:genetics*; D008253:Macaca mulatta; D016366:Open Reading Frames; D010957:Plasmids / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000235:genetics
D018076:DNA, Complementary; D011993:Recombinant Fusion Proteins