Baculoviral transduction of primary fibroblasts

Baculoviruses are double-stranded DNA viruses that infect insects. Baculoviruses have been shown to efficiently transduce a number of different cell lines in vitro, however cultured primary cells have proven difficult to transduce. During this study, the optimal parameters for baculoviral transduction of cultured primary fibroblasts were determined. Transduced cells were identified by the expression of the enhanced green fluorescent protein under control of the SV40 promoter. During baculoviral infection, two types of virions are produced, the Occlusion Derived Virus (ODV) and Budded Virus (BV). The major envelope glycoprotein of BV is gp64, which plays an essential role in viral entry into target insect cells by membrane fusion. Short cationic peptides have been shown to bind to the cell surface of many cell types. Polyarginine stretched between 5 and 15 residues have proven to be most effective. A polyarginine motif was designed for insertion into the native gp64 sequence in an attempt to target the cell membrane of primary fibroblasts more efficiently. The transduction efficiency of this mutant virus was then compared to the efficiency of a non-mutated control virus. The use of this mutant baculovirus takes advantage of the efficiency of viral gene delivery, while avoiding the problems of viral replication or integration that are posed by adenoviral and retroviral vectors respectively