New vectors for the construction of double recombinant adenoviruses.

A set of plasmids designed for the construction of recombinant adenoviral vectors is described, which contain two expression cassettes, one in the early region 1 (E1) and the other in the early region 3 (E3). Two cloning steps in E. coli and a transfection of the resulting cosmid into 293 cells are sufficient to recover the recombinant virus. The method has been optimised to facilitate the introduction of the genes of interest in their respective regions and the reconstitution of the entire sequence of the recombinant adenoviral DNA in E. coli. The vectors are easy to handle and generate homogenous virus preparations. To illustrate the efficiency of the method, an adenovirus was constructed expressing the E. coli beta-galactosidase deltaM15 mutant in the E1 region and the complementing lacZ alpha-peptide in the E3 region.