Cell-based reporter assay for screening inhibitors of coronavirus RNA-dependent RNA polymerase activity

A screening composition for a therapeutic agent for coronavirus infection, comprising:

• (a) a CoV RdRp expression vector comprising a FLAG-labeled CoV (coronavirus) nsp12 gene at the N-terminus;
• and
• (b) a bicistronic reporter vector comprising:
• a firefly luciferase gene in the sense direction ((+)FLuc) and a NanoLuc luciferase gene in the antisense direction ((−)NLuc);
• wherein the (−)NLuc forms domains (3′-UTR, NLuc and 5′-UTR) interposed between 3′-UTR and 5′-UTR in the antisense direction derived from CoV;
• wherein the 3′-UTR, NLuc, and 5′-UTR domains in the antisense direction are in the form in which a ribozyme self-cleaving sequence derived from hepatitis delta virus (HDV) is bound to each of the 5′- and 3′-ends thereof;
• and a firefly luciferase gene in the sense direction independently of the domain in the antisense direction.