Generation of Recombinant MVA Expressing SARS-CoV-2 RBD

For the generation of recombinant MVA expressing SARS-CoV-2 RBD wt and RBD M7 the shuttle vectors pMVA Trans TK-SARS-2 RBD wt and pMVA Trans TK SARS-2 RBD M7 were cloned. The MVA shuttle vectors were designed in a way that the antigens SARS-CoV-2 RBD wt and SARS-CoV-2 RBD M7 antigens can be inserted into the thymidine kinase (TK) locus J2R of the parental virus MVA CR19 TK-GFP under the transcriptional control of the early/late modified H5 promoter (mH5) via homologous recombination. The MVA shuttle vectors also include the reporter gene β-galactosidase (β-Gal) between the two left arm sequences of the TK locus for screening of recombinant MVAs. After several plaque purification rounds the reporter gene is lost by internal homologous recombination events resulting in a pure (reporter-free) recombinant MVA.