FIGURE 1.

Expression yields increase by codon optimization and sfGFP fusion. (a) HA expression plasmids: Schematic representation of the HA expression cassette used. The HA ectodomain encoding sequence, under the control of CMV, was cloned in frame with DNA sequences coding for the CD5 signal peptide. C‐terminally we cloned GCN4 trimerization domain and a TEV cleavable Strep‐tag II in which we inserted a sfGFP between the TEV site and the Strep‐tag. Standard restriction sites for cloning are indicated. (b) Expression of A/TX/12, A/CH/13 H3N2, and A/MI/15 H1N1: Supernatants were analyzed by SDS–PAGE followed by western blotting, recombinant proteins were detected using a mouse HRP labelled anti‐Strep‐tag antibody. (c) Quantification: sfGFP emission was directly measured in the supernatant. Shown is a representative of four independent expression experiments over a time span of 4 weeks. (d) Avian coronavirus M41 receptor binding domain fused with sfGFP: Supernatants were analyzed by SDS–PAGE followed by western blotting, where recombinant proteins were detected using a mouse anti‐Strep‐tag antibody