Co-expression Of GPV-VP3 Gene And Goose Interferon ? Gene In Recombinant Fowlpox Virus

Goose Parvovirusis(GP),commonly known as goose plague and could be found around the world,is an acute,subacute and septic infectious disease caused by goose parvovirus(GPV).With the extremely high occurence and mortality,GP brings huge economic loss to goose industry.At present,the prevention and treatment of GP mainly depend on attenuated vaccination on mother goose to immunize the young goose.Although the application can efficiently control the spread of GP,its risk of spreading the poison and the possibility of virulence recovering has greatly limited the use of the vaccine.Therefore,there is an urgent need to develop a safe and effective new vaccine.Recombinant poxvirus vaccines are appreciated by many researchers for their advantages such as the safety,stability and ability to express exogenous genes.In particular,recombinant fowlpox virus(FPV)widely used,because with replicative non-essential areas,it can be inserted into foreign genes,and continually express exogenous genes,stimulating the body to continue to produce specific antibodies.GPV genome mainly encodes three structural proteins,namely,VP1,VP2,VP3,where VP3 is the main capsid protein of the virus,and is capable of stimulating the body to produce neutralizing antibody.Interferon(IFN?)can enhance the body's anti-virus ability by raising the level of cellular immunity.A large number of studies have shown that it is a good immune adjuvant.In this study,we construct a recombinant fowlpox virus co-expressing the GPV-VP3 gene and the goose IFN? gene.The pMD-18T-GPV-IFN? plasmid is used as a template and PCR is performed with the primer containing EcoR I restriction site.The target gene and reporter gene are inserted into the transfer vector pSY681 to obtain the LP2EP2-VP3-LacZ gene,and the pSY538-?-LacZ gene is inserted into the pSY538 vector promoter.The LP2EP2-VP3-LacZ gene is cloned into the Not I restriction site of pSY681-LP2EP2-IFN? by Clon Express II One Step Cloning Kit to complete the construction of GPV-VP3 gene and goose IFN? gene transfer vector(pSY-GoIFN?-VP3)(CEF)is transfected into chicken embryo fibroblasts(CEF)by lipofectamine method.The recombinant virus is screened by blue spot method and subjected to 8 rounds of plaque purification,as well as PCR identification and indirect immunofluorescence(IFA)analysis.The results show that the recombinant virus rFPV-GoIFN?-VP3 could stably express the foreign gene.This study lays the foundation for the further development of rFPV-GoIFN?-VP3 recombinant fowlpox virus vaccine,providing reference for the development of recombinant fowlpox virus vaccine for other avain infectious diseases.