| Duck viral hepatitis (DVH) caused by duck hepatitis virus (DHV) is a highly mortal disease andspreads rapidly in four-week-old ducklings characterized by high mortality and liver lesions withhemorrhagic foci. Three distinct types of DHV have been identified from diseased ducklings (Woolcock,2003; Monroe et al.,2005). DHV-I is prevalent in China, which lead large economic loss for duckery.Currently, control of the disease is based upon the use of classic vaccine. Thus, to develop safer andmore effective and newer vaccine to protect and control this disease is a promising direction in thefuture.VP1is likely to be the most important structural protein, being the most external and dominant ofall surface proteins and neutralizing monoclonal antibodies (Feigelstock et al.,1996). In this study, theVP1gene was amplified by RT-PCR, and cloned into transfer vector pShuttle-CMV. Aftertransformation of Pme I-linearized recombinant plasmid pShuttle-CMV-VP1into Escherichia colibacteria strain BJ5183containing the vector pAdEasy-1, recombinant plasmid containing VP1proteingene (pAd-VP1) was obtained and identified with PCR and sequencing. Upon transfection of PacⅠ-linearized plasmid pAd-VP1in AD293cell line, a recombinant adenovirus was obtained and named asrAd-VP1. rAd-VP1was purified by small cesium choride density centrifugation kit, the titer wasdetermined to be6.3×107TCID50/ml. The expression of the VP1protein in the AD293cells infectedwith rAd-VP1was confirmed with monoclonal antibody to DHV-VP1by Western blotting and IFA.7days ducklings were inoculated with5*107TCID50rAd-VP1, and generated antibody against HP-1detected by indirect ELISA and protect ducks against HP-1. It indicated that the rAd-VP1was able tocreate special IgG against the DHV-I in duck.The recombinant adenovirus expressing VP1protein of DHV-I was successfully constructed andinduced special humoral immunity against DHV-I. The rAd-VP1laid the foundation for creating thenew vaccine. |
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