:: Volume 7, Issue 1 And 2 (7-2013) ::
Iran J Virol 2013, 7(1 And 2): 15-20 Back to browse issues page
Construction of Recombinant Bacmid DNA Encoding Newcastle Disease Virus (NDV) Fusion Protein Gene
Mohammad reza Shafaati *, Majid Moghbeli, Roholah Dorostkar
Department of Microbiology, Islamic Azad University, Damaghan Branch, Damaghan, Iran
Abstract:   (5539 Views)
Background and Aims: Newcastle disease virus (NDV) is one of the major pathogen in poultry. Vaccination is intended to control the disease as an effective solution nevertheless this virus is a growing threat to the poultry industry. F gene open reading frame (ORF) from NDV is 1650 bp, encoding a protein of 553 amino acids that can induce protective immunity alone. The F glycoprotein on the surface of NDV is important for virus infectivity and pathogenicity. Towards protection goal, the full-length of F gene was isolated using specific primers and cloned into the baculovirus derived bacmid shuttle vector to produce recombinant F-protein in insect cells. Materials and Methods: F gene ORF from avirulent strain La Sota of NDV was amplified by RT-PCR to produce F cDNA. The amplicon was cloned firstly into the T/A cloning vector and then subcloned into the pFastBac Dual donor plasmid through NcoI/KpnI sites. After the verification of cloning process by PCR and enzymatic digestion analysis, the accuracy of F gene ORF in the T/A cloning vector was confirmed by sequencing. Finally, F-containing recombinant bacmid was subsequently generated and confirmed by PCR using F specific primers and M13 universal primers. Results: Results showed that a recombinant baculovirus containing a correct and in framework sequence of Newcastle F gene under the control of p10 promoter was constructed. Conclusion: The above mentioned F-containing recombinant baculovirus, in addition to its independent application, can be used with other individual recombinant baculoviruses expressing NH and NP genes to produce Newcastle VLPs in insect cell line.
Keywords: Newcastle disease, F protein, Baculovirus, Bacmid shuttle vector
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Type of Study: Original article | Subject: General
Received: 2014/12/9 | Accepted: 2014/12/9 | Published: 2014/12/9

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Volume 7, Issue 1 And 2 (7-2013) Back to browse issues page