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http://conacyt.repositorioinstitucional.mx/jspui/handle/1000/8068| Rapid cloning-free mutagenesis of new SARS-CoV-2 variants using a novel reverse genetics platform | |
| Fabian Otte Enja Tatjana Kipfer David Hauser Martin Joseph LETT Lorena Urda Yuepeng Zhang Christopher Lang mohamed chami Christian Mittelholzer Thomas Klimkait | |
| Acceso Abierto | |
| Atribución-NoComercial-SinDerivadas | |
| https://doi.org/10.1101/2023.05.11.540343 | |
| https://www.biorxiv.org/content/10.1101/2023.05.11.540343v1 | |
| Abstract Reverse genetic systems enable engineering of RNA virus genomes and are instrumental to study RNA virus biology. With the recent outbreak of the COVID-19 pandemic, already established methods were challenged by the large genome of SARS-CoV-2. Herein we present an elaborated strategy for the rapid and straightforward rescue of recombinant plus-stranded RNA-viruses with high sequence fidelity, using the example of SARS-CoV-2. The strategy called CLEVER (CLoning-free and Exchangeable system for Virus Engineering and Rescue) is based on the intracellular recombination of transfected overlapping DNA fragments allowing the direct mutagenesis within the initial PCR-amplification step. Furthermore, by introducing a linker fragment (harboring all heterologous sequences) viral RNA can directly serve as template for manipulation and rescue of recombinant mutant virus, without any cloning-step needed. Overall, this strategy will facilitate recombinant SARS-CoV-2 rescue and accelerate its manipulation. Using our protocol, newly emerging variants can quickly be engineered to further elucidate its biology. | |
| bioRxiv | |
| 12-05-2023 | |
| Preimpreso | |
| Inglés | |
| Público en general | |
| VIRUS RESPIRATORIOS | |
| Aparece en las colecciones: | Materiales de Consulta y Comunicados Técnicos |
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| Rapid cloning-free mutagenesis of new SARS-CoV-2 variants using a novel reverse genetics platform.pdf | 4.12 MB | Adobe PDF | Visualizar/Abrir |