Recombinant Fowlpox Virus Expressing F of Newcastle Disease Virus and HA of Subtype H9N2 of Avian Influenza Virus
共表达新城疫病毒F基因和H9亚型禽流感病毒HA基因的重组鸡痘病毒

The fusion protein (F) gene of Newcastle disease virus strain F48E8 was obtained by PCR and its sequence was determined. To form recombinant plasmid p7SHF, the F gene in pGEM-TF and the hemagglutinin (HA) gene of AIV, A/Chicken/China/F/1998(H9N2) in pUCHA were cut and inserted into the randomly selected nonessential regions of the fowlpox virus (FPV) under the control of fowlpox virus promoter P E/L and the synthetic promoter Ps respectively. Then the P11-LacZ reporter gene from pppG18 was cloned into p7SHF resulting in recombinant transferring vector pFPVHF, which was transfected into chicken embryo fibroblast (CEF) monolayers pre-infected with Chinese vaccine strain 282E4 of FPV to generate recombinant fowlpox virus co-expressing HA and F (rFPV-HA-F). By selection of blue plaques on the CEF overlaid with agar containing X-gal, rFPV-HF-F was screened and purified. PCR and southern blot assays indicated that the F and HA genes had been inserted into the genomic DNA of FPV. Subsequently expression of F and HA in CEF infected with rFPV-HA-F was confirmed by indirect immunofluorescence assay.