Cloning, expression, purification and characterization of UL24 partial protein of duck enteritis virus.
The UL24 gene of duck enteritis virus (DEV) is conserved across herpesviruses, but its protein characterization has not been reported. We expressed the UL24 gene in Escherichia coli BL21 from a recombinant plasmid pET32a/DEV-UL24 and used the resulting protein to raise antiserum. This antiserum recognized a 38-kDa protein in lysates from infected cells. SDS-PAGE analysis showed that the UL24 partial protein was highly expressed after induction by 0.4 mM IPTG at 30 degrees for 6 h. The results of purification revealed that expression protein was more purified using the method of electrophoresis than that of chromatography, but the yield was lower. In immunogenicity analysis, the protein could significantly elicit a specific antibody response in immunized ducklings when compared with the control groups, and the titers against expression protein reached the peak 1:5,120 (OD(450nm) = 2.5) on day 28 after immunization, while with mean titers of 1:10,240 (OD(450nm) = 3.37) in DEV commercial attenuated vaccine strain immunized duckling groups. It showed that expression protein is immunogenic in laboratory ducklings. On the basis of subcellular location, UL24 appeared to be predominantly nuclear membrane-associated, especially at later times in infection, and provided a good tool to further study the biofunctions of UL24 protein.
Jia
Renyong
R
Avian Disease Research Center, College of Veterinary Medicine of Sichuan Agricultural University, Ya'an 625014, China.
Cheng
Anchun
A
Wang
Mingshu
M
Zhu
Dekang
D
Ge
Han
H
Xin
Hongyi
H
Liu
Fei
F
Luo
Qihui
Q
Guo
Yufei
Y
Qi
Xuefeng
X
Yin
Zhongqiong
Z
Chen
Xiaoyue
X
D000818:Animals; D000914:Antibodies, Viral / Q000096:biosynthesis / Q000276:immunology; D001715:Bird Diseases / Q000276:immunology / Q000517:prevention & control / Q000821:virology*; D003001:Cloning, Molecular; D004372:Ducks; D004591:Electrophoresis, Polyacrylamide Gel; D004751:Enteritis / Q000517:prevention & control / Q000662:veterinary* / Q000821:virology; D015870:Gene Expression; D006564:Herpesviridae / Q000235:genetics* / Q000276:immunology / Q000378:metabolism; D006566:Herpesviridae Infections / Q000276:immunology / Q000378:metabolism / Q000517:prevention & control / Q000662:veterinary*; D007106:Immune Sera / Q000276:immunology; D016233:Immunotherapy, Active; D009687:Nuclear Proteins / Q000096:biosynthesis / Q000276:immunology / Q000302:isolation & purification; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000276:immunology / Q000302:isolation & purification; D014764:Viral Proteins / Q000302:isolation & purification / Q000502:physiology
D000914:Antibodies, Viral; D007106:Immune Sera; D009687:Nuclear Proteins; D011994:Recombinant Proteins; D014764:Viral Proteins