Precursor firefly and Gaussia luciferase reporter gene constructs were generated by GenScript. These constructs have the following schematic make up: hepatitis delta virus (HDV) ribozyme -AloI restriction site -firefly or Gaussia luciferase gene -BaeI restriction site -human PolI promoter. These constructs did not yet contain IAV 59 and 39 UTR sequences. 39 UTR sequences were introduced by ligation of primer dimers into the AloI-digested constructs. Subsequently, primer dimers corresponding to 59 UTR sequences were ligated into the resulting BaeI-digested plasmid. The AloI and BaeI restriction recognition sites are completely removed by this procedure and the corresponding PolI transcripts are similar to genuine vRNA segments with the exception of the coding region. In addition, we generated an extended version of the Gaussia luciferase reporter gene construct by introduction of a firefly luciferase gene fragment. As a result, the length of the extended Gaussia luciferase vRNA is identical to that of the corresponding firefly luciferase-encoding vRNA. To generate this construct, a NotI-digested PCR fragment, corresponding to part of the firefly luciferase gene (nucleotide 559 -stop codon) and containing flanking NotI restriction sites, was ligated into the NotIrestricted precursor Gaussia luciferase plasmid, immediately downstream of the Gaussia luciferase gene stop codon. The IAV segment UTRs were inserted in the extended Gaussia luciferase plasmid in the same manner as described above. For a schematic overview of the reporter constructs, see Figure 1A .