Virus-like particle (VLP)-based vaccines represent a promising approach for production of new generation vaccine with improved safety and efficacy. In order to construct a VLP-based NH5NI vaccine
herein we constructed a baculovirus transfer vector, which is able to support the independent expression of different open reading frames (ORFs), carrying a gene cassette containing hemaglutinin sequence trom the strain A/chicken/hatay/2004(H5NI). The gene encoding hemagglutinin was amplified by PCR using a specific primer set PFSP-F and HApBac-R which has initiation codon, Kozak sequence, and BamHI site at 5' of forward primer and HindIII site at 5' of reverse primer. The PCR product (HA gene cassette) was cloned into pCR2.l vector and sequenced. The HA gene cassette was then excised trom recombinant pCR2.1 vector by BamHI and HindIII and inserted into the baculovirus transfer vector (pBluBac4.5/V5-His-TOPO) previously digested by the same restriction enzymes to yield recombinant pBluBacHA plasmid. The structure and nucleotide sequences of HA gene cassette in pBluBacHA plasmid was confirmed by restriction enzyme analysis and DNA sequencing. The correct clones, in the next step, will be co-transfected with DNA of baculovirus into Spodopterafrugiperda (Sf9) cells to generate H5Nl VLP - based vaccine.