title: Ad 2.0: a novel recombineering platform for high-throughput generation of tailored adenoviruses.
abstract: Recombinant adenoviruses containing a double-stranded DNA genome of 26-45 kb were broadly explored in basic virology, for vaccination purposes, for treatment of tumors based on oncolytic virotherapy, or simply as a tool for efficient gene transfer. However, the majority of recombinant adenoviral vectors (AdVs) is based on a small fraction of adenovirus types and their genetic modification. Recombineering techniques provide powerful tools for arbitrary engineering of recombinant DNA. Here, we adopted a seamless recombineering technology for high-throughput and arbitrary genetic engineering of recombinant adenoviral DNA molecules. Our cloning platform which also includes a novel recombination pipeline is based on bacterial artificial chromosomes (BACs). It enables generation of novel recombinant adenoviruses from different sources and switching between commonly used early generation AdVs and the last generation high-capacity AdVs lacking all viral coding sequences making them attractive candidates for clinical use. In combination with a novel recombination pipeline allowing cloning of AdVs containing large and complex transgenes and the possibility to generate arbitrary chimeric capsid-modified adenoviruses, these techniques allow generation of tailored AdVs with distinct features. Our technologies will pave the way toward broader applications of AdVs in molecular medicine including gene therapy and vaccination studies.
authors.affiliation: Max von Pettenkofer-Institute, Department of Virology, Ludwig-Maximilians-University Munich, Munich, Germany.
authors.forename: Martin
authors.initials: M
authors.lastname: Mück-Häusl
authors.affiliation: Institute of Virology and Microbiology, Center for Biomedical Education and Research (ZBAF), Department of Human Medicine, Faculty of Health, University of Witten/Herdecke, Witten, Germany.
authors.forename: Manish
authors.initials: M
authors.lastname: Solanki
authors.affiliation: Institute of Virology and Microbiology, Center for Biomedical Education and Research (ZBAF), Department of Human Medicine, Faculty of Health, University of Witten/Herdecke, Witten, Germany.
authors.forename: Wenli
authors.initials: W
authors.lastname: Zhang
authors.affiliation: Max von Pettenkofer-Institute, Department of Virology, Ludwig-Maximilians-University Munich, Munich, Germany Institute of Virology, Albert-Ludwigs-University Freiburg, Freiburg, Germany.
authors.forename: Zsolt
authors.initials: Z
authors.lastname: Ruzsics
authors.affiliation: Max von Pettenkofer-Institute, Department of Virology, Ludwig-Maximilians-University Munich, Munich, Germany Institute of Virology and Microbiology, Center for Biomedical Education and Research (ZBAF), Department of Human Medicine, Faculty of Health, University of Witten/Herdecke, Witten, Germany anja.ehrhardt@uni-wh.de.
authors.forename: Anja
authors.initials: A
authors.lastname: Ehrhardt
mesh_terms: D000256:Adenoviridae / Q000235:genetics*; D022202:Chromosomes, Artificial, Bacterial; D003001:Cloning, Molecular / Q000379:methods*; D004279:DNA, Viral; D005818:Genetic Engineering; D005822:Genetic Vectors; D057809:HEK293 Cells; D006801:Humans; D011995:Recombination, Genetic
chemical_list: D004279:DNA, Viral