Advanced Fusion Strategies for the Production of Functionalized Potato Virus X Virions

Plant virions are ideal for nanotechnology applications because they are structurally diverse and can self-assemble naturally, allowing for large-scale production in plants by molecular farming. Potato virus X (PVX) is particularly amenable due to the unique properties of its filamentous and flexible capsid, but efficient strategies are required to adapt the surface properties of PVX, such as the attachment of proteins and peptides. This chapter describes the selection and utilization of 2A ribosomal skip sequences, allowing the presentation of heterologous proteins and peptides as N-terminal fusions to the PVX coat protein at different densities. Another strategy for the rapid modification of PVX capsids is the plug-and-display module of the SpyTag/SpyCatcher system. The SpyTag can be presented on the PVX surface, allowing for the attachment of any protein fused to the SpyCatcher sequence.