In vitro cloning of canine parvovirus NS1 gene and reporter gene GFP in eukaryotic expression vector pVIVO2-mcs and characterization of the double gene construct in mammalian cells

The use of chemotherapy and/or radiotherapy for treatment of cancer is limited due to genotoxic side effects on healthy cells, involvement of anti-apoptotic signal transduction pathways that prevent cell death, and requirement of functional p53 for induction of apoptosis in cancerous cells. Efforts are beings made worldwide to develop new anticancer therapies as an alternative to chemotherapy. And viral gene therapy is one of the most potent therapeutics that is being ventured worldwide. Canine parvovirus-2 (CPV-2) is one of those viruses that have an inherent oncolytic property. The non-structural protein-1 (NS1 protein) of CPV-2 plays a major role in parvoviral cytotoxicity and pathogenicity in permissive cells. The oncolytic potential of CPV2-NS1 has been established in vitro. Prior to taking up the in vivo studies, the present study was undertaken to clone Canine Parvovirus NS1 gene and reporter gene GFP in eukaryotic expression vector pVIVO2-mcs, and to characterize the double construct in mammalian cells. The genes were successfully cloned in pVIVO2-mcs and characterized for their expression as demonstrated by fluorescence microscopy and immunofluorescence staining. This characterized double gene construct will further be used to evaluate the oncolytic potential of CPV-2 NS1 in experimentally induced in vivo tumour model.