Seamless assembly of recombinant adenoviral genomes from high-copy plasmids.
The adenoviruses are essential tools for basic research and therapeutic development. Robust methods for the generation of mutant and recombinant viruses are crucial for these diverse applications. Here we describe a simple plasmid-based method that permits highly efficient modification of the adenov...
| Main Authors: | , , |
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| Format: | Article |
| Language: | English |
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Public Library of Science (PLoS)
2018-01-01
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| Series: | PLoS ONE |
| Online Access: | http://europepmc.org/articles/PMC6021080?pdf=render |
| _version_ | 1811320437300264960 |
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| author | Jessica J Miciak Jason Hirshberg Fred Bunz |
| author_facet | Jessica J Miciak Jason Hirshberg Fred Bunz |
| author_sort | Jessica J Miciak |
| collection | DOAJ |
| description | The adenoviruses are essential tools for basic research and therapeutic development. Robust methods for the generation of mutant and recombinant viruses are crucial for these diverse applications. Here we describe a simple plasmid-based method that permits highly efficient modification of the adenoviral genome and rapid production of high-titer virus stocks. The 36-kilobase genome of adenovirus serotype 5 was divided into seven tractable blocks that could be individually modified in a single step and reassembled in vitro. Because the system is composed of compact modules, modifications at different loci can be readily recombined. Viral assemblies were delivered to packaging cells by electroporation, a strategy that consistently resulted in the de novo production of 108 infectious units in 3-5 days. In principle, a similar strategy could be applied to any other adenovirus serotype or to other double-strand DNA viruses. |
| first_indexed | 2024-04-13T12:59:27Z |
| format | Article |
| id | doaj.art-4387aeec482f4aa9905c6d66346004db |
| institution | Directory Open Access Journal |
| issn | 1932-6203 |
| language | English |
| last_indexed | 2024-04-13T12:59:27Z |
| publishDate | 2018-01-01 |
| publisher | Public Library of Science (PLoS) |
| record_format | Article |
| series | PLoS ONE |
| spelling | doaj.art-4387aeec482f4aa9905c6d66346004db2022-12-22T02:45:57ZengPublic Library of Science (PLoS)PLoS ONE1932-62032018-01-01136e019956310.1371/journal.pone.0199563Seamless assembly of recombinant adenoviral genomes from high-copy plasmids.Jessica J MiciakJason HirshbergFred BunzThe adenoviruses are essential tools for basic research and therapeutic development. Robust methods for the generation of mutant and recombinant viruses are crucial for these diverse applications. Here we describe a simple plasmid-based method that permits highly efficient modification of the adenoviral genome and rapid production of high-titer virus stocks. The 36-kilobase genome of adenovirus serotype 5 was divided into seven tractable blocks that could be individually modified in a single step and reassembled in vitro. Because the system is composed of compact modules, modifications at different loci can be readily recombined. Viral assemblies were delivered to packaging cells by electroporation, a strategy that consistently resulted in the de novo production of 108 infectious units in 3-5 days. In principle, a similar strategy could be applied to any other adenovirus serotype or to other double-strand DNA viruses.http://europepmc.org/articles/PMC6021080?pdf=render |
| spellingShingle | Jessica J Miciak Jason Hirshberg Fred Bunz Seamless assembly of recombinant adenoviral genomes from high-copy plasmids. PLoS ONE |
| title | Seamless assembly of recombinant adenoviral genomes from high-copy plasmids. |
| title_full | Seamless assembly of recombinant adenoviral genomes from high-copy plasmids. |
| title_fullStr | Seamless assembly of recombinant adenoviral genomes from high-copy plasmids. |
| title_full_unstemmed | Seamless assembly of recombinant adenoviral genomes from high-copy plasmids. |
| title_short | Seamless assembly of recombinant adenoviral genomes from high-copy plasmids. |
| title_sort | seamless assembly of recombinant adenoviral genomes from high copy plasmids |
| url | http://europepmc.org/articles/PMC6021080?pdf=render |
| work_keys_str_mv | AT jessicajmiciak seamlessassemblyofrecombinantadenoviralgenomesfromhighcopyplasmids AT jasonhirshberg seamlessassemblyofrecombinantadenoviralgenomesfromhighcopyplasmids AT fredbunz seamlessassemblyofrecombinantadenoviralgenomesfromhighcopyplasmids |