This article presents a simplified method for generating recombinant adenoviruses, which are widely used in gene expression studies and therapeutic applications. The authors describe a strategy that minimizes enzymatic manipulations and uses homologous recombination in bacteria rather than eukaryotic cells. The process involves creating a recombinant adenoviral plasmid with a minimum of enzymatic steps, followed by transfections into mammalian packaging cell lines. Green fluorescent protein (GFP) is incorporated into the viral backbone to facilitate the monitoring of transfection and infection efficiency. This approach allows for the production of homogeneous viruses without the need for plaque purification, significantly reducing the time and effort required for viral production. The system is versatile, allowing for the inclusion of up to 10 kb of transgene sequences and multiple transgenes from a single virus. The authors also detail the construction of various vectors and the optimization of the recombination process, demonstrating the efficiency and practical advantages of their method.This article presents a simplified method for generating recombinant adenoviruses, which are widely used in gene expression studies and therapeutic applications. The authors describe a strategy that minimizes enzymatic manipulations and uses homologous recombination in bacteria rather than eukaryotic cells. The process involves creating a recombinant adenoviral plasmid with a minimum of enzymatic steps, followed by transfections into mammalian packaging cell lines. Green fluorescent protein (GFP) is incorporated into the viral backbone to facilitate the monitoring of transfection and infection efficiency. This approach allows for the production of homogeneous viruses without the need for plaque purification, significantly reducing the time and effort required for viral production. The system is versatile, allowing for the inclusion of up to 10 kb of transgene sequences and multiple transgenes from a single virus. The authors also detail the construction of various vectors and the optimization of the recombination process, demonstrating the efficiency and practical advantages of their method.