Rapid reconstruction of SARS-CoV-2 using a synthetic genomics platform
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Date
2020-05-04Author
Thi Nhu Thao, Tran
Labroussaa, Fabien
Ebert, Nadine
V’kovski, Philip
Stalder, Hanspeter
Portmann, Jasmine
Kelly, Jenna
Steiner, Silvio
Holwerda, Melle
Kratzel, Annika
Gultom, Mitra
Schmied, Kimberly
Laloli, Laura
Hüsser, Linda
Wider, Manon
Pfaender, Stephanie
Hirt, Dagny
Cippà, Valentina
Crespo-Pomar, Silvia
Schröder, Simon
Muth, Doreen
Niemeyer, Daniela
Corman, Victor M.
Müller, Marcel A.
Drosten, Christian
Dijkman, Ronald
Jores, Joerg
Thiel, Volker
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Summary in foreign language
Reverse genetics has been an indispensable tool to gain insights into viral pathogenesis and vaccine development. The genomes of large RNA viruses, such as those from coronaviruses, are cumbersome to clone and manipulate in Escherichia coli owing to the size and occasional instability of the genome1,2,3. Therefore, an alternative rapid and robust reverse-genetics platform for RNA viruses would benefit the research community. Here we show the full functionality of a yeast-based synthetic genomics platform to genetically reconstruct diverse RNA viruses, including members of the Coronaviridae, Flaviviridae and Pneumoviridae families. Viral subgenomic fragments were generated using viral isolates, cloned viral DNA, clinical samples or synthetic DNA, and these fragments were then reassembled in one step in Saccharomyces cerevisiae using transformation-associated recombination cloning to maintain the genome as a yeast artificial chromosome. T7 RNA polymerase was then used to generate infectious RNA to rescue viable virus. Using this platform, we were able to engineer and generate chemically synthesized clones of the virus, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)4, which has caused the recent pandemic of coronavirus disease (COVID-19), in only a week after receipt of the synthetic DNA fragments. The technical advance that we describe here facilitates rapid responses to emerging viruses as it enables the real-time generation and functional characterization of evolving RNA virus variants during an outbreak.
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SARS-CoV-2Link to resource
https://www.nature.com/articles/s41586-020-2294-9Collections
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