A protocol for rapid generation of recombinant adenoviruses using the AdEasy system

Nat Protoc. 2007;2(5):1236-47. doi: 10.1038/nprot.2007.135.


Recombinant adenoviruses provide a versatile system for gene expression studies and therapeutic applications. We have developed an approach that simplifies the generation and production of such viruses called the AdEasy system. A recombinant adenoviral plasmid is generated with a minimum of enzymatic manipulations, employing homologous recombination in bacteria rather than in eukaryotic cells. After transfection of such plasmids into a mammalian packaging cell line, viral production is conveniently followed with the aid of GFP encoded by a gene incorporated into the viral backbone. This system has expedited the process of generating and testing recombinant adenoviruses for a variety of purposes. In this protocol, we describe the practical aspects of using the AdEasy system for generating recombinant adenoviruses. The full protocol usually takes 4-5 weeks to complete.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adenoviridae / genetics*
  • Bacteria
  • Cells, Cultured
  • Genetic Engineering / methods*
  • Genetic Vectors / genetics*
  • Green Fluorescent Proteins / genetics
  • Humans
  • Recombination, Genetic*
  • Transfection / methods*


  • Green Fluorescent Proteins