A simplified system for generating recombinant adenoviruses

TLDR

Recombinant adenoviruses provide a versatile system for gene expression studies and therapeutic applications. We report a strategy that simplifies the generation and production of such viruses. The method uses bacterial homologous recombination to create a plasmid with minimal enzymatic steps, then transfects it into mammalian packaging cells where viral production is monitored via GFP expression. The procedure yields homogeneous viruses without plaque purification and accelerates generation and testing of recombinant adenoviruses for diverse applications.

Abstract

Recombinant adenoviruses provide a versatile system for gene expression studies and therapeutic applications. We report herein a strategy that simplifies the generation and production of such viruses. A recombinant adenoviral plasmid is generated with a minimum of enzymatic manipulations, using homologous recombination in bacteria rather than in eukaryotic cells. After transfections of such plasmids into a mammalian packaging cell line, viral production is conveniently followed with the aid of green fluorescent protein, encoded by a gene incorporated into the viral backbone. Homogeneous viruses can be obtained from this procedure without plaque purification. This system should expedite the process of generating and testing recombinant adenoviruses for a variety of purposes.