Rapid construction of adenoviral vectors by lambda phage genetics

Journal of Virology
Duncan McVeyImre Kovesdi

Abstract

Continued improvements of adenoviral vectors require the investigation of novel genome configurations. Since adenovirus can be generated directly by transfecting packaging cell lines with viral genomes isolated from plasmid DNA, it is possible to separate genome construction from virus production. In this way failure to generate a virus is not associated with an inability to generate the desired genome. We have developed a novel lambda-based system that allows rapid modification of the viral genome by double homologous recombination in Escherichia coli. The recombination reaction and newly generated genome may reside in a recombination-deficient bacterial host for enhanced plasmid stability. Furthermore, the process is independent of any restriction endonucleases. The strategy relies on four main steps: (i) homologous recombination between an adenovirus cosmid and a donor plasmid (the donor plasmid carries the desired modification[s] and flanking regions of homology to direct its recombination into the viral genome); (ii) in vivo packaging of the recombinant adenoviral cosmids during a productive lambda infection; (iii) transducing a recombination-deficient E. coli lambda lysogen with the generated lysate (the lysogen inhibits ...Continue Reading

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Citations

Jul 27, 2007·Molecular Biotechnology·Jason G D GallDouglas E Brough
Oct 21, 2010·Virology Journal·Duncan McVeyJason G D Gall
Aug 5, 2015·Journal of Virological Methods·Yanlong PeiÉva Nagy
Jan 26, 2007·Vaccine·Franck LemialeJason G D Gall
Apr 21, 2010·The Journal of Cell Biology·Kohei YamamizuJun K Yamashita
Dec 4, 2003·The Journal of General Virology·Duncan McVeyImre Kovesdi
Oct 30, 2004·The Journal of Gene Medicine·Gunhild M MaelandsmoRobin J Parks
Oct 31, 2008·Current Protocols in Neuroscience·Thomas SouthgateMaria G Castro
Sep 19, 2014·Clinical and Vaccine Immunology : CVI·Benjamin J KoestlerChristopher M Waters
Jun 1, 2009·Viruses·Kenneth Lundstrom

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