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Construction and Identification of the Recombinant Goatpox Virus with ORF8—ORF18 Deleted

ZHENG Min1,2,LIU Qi1,JIN Ning-yi2,SHI Kai-chuang1,LIANG Yuan1,MO Sheng-lan1,QU Su-jie1,LU Wen-jun1,HU Bo2(1.Guangxi Center for Animal Disease Control and Prevention,Nanning 530001,China;2.TheMilitary Veterinary Institute,Academy of Military Medical Sciences,Changchun 130062,China)  
This experiment was conducted to develop a safer,high effective live goatpox virus(GTPV) attenuated vaccine and viral vector.Genome fragments of GTPV AV41 strain,including ORF7—ORF8 and ORF18—ORF19,were cloned by PCR as flanking sequences for homologous recombinant,respectively.The transfer shuttle plasmid pCF-Eg was constructed by inserting the expression cassettes of EGFP gene and gpt gene.The pCF-Eg and GTPV AV41 were co-transfected into LT cells with lipofectin,and recombinant virus was selected by fluorescence and PCR with EGFP gene primer respectively.Then,resulting recombinant virus was examined by fluorescence,PCR and TCID50 during passaging to 10th in LT cells.A recombinant GTPV with ORF8—ORF18 deleted was obtained and named as vCF-Eg.It was stable,and shared same multiplication ability with its parental virus(AV41) in LT cells.It suggests that genomic region of ORF8—ORF18 may be non-essential regions for replication.
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