Many complex recombinant proteins require oxidative folding, the assembly of subunits, and post-translational modifications such as N-glycosylation. These features are not provided by most microbial cells so higher eukaryotic hosts are required. Mammalian cells are widely used for this purpose but both the development of stable mammalian cell lines and the more rapid transient expression of recombinant proteins in animal cell lines require special infrastructure and expertise as well as expensive media. Therefore, transient expression in plants is an attractive alternative for the production of recombinant proteins in low microgram to higher milligram amounts.
The procedure includes the use of pre-grown wild-type Nicotiana benthamiana plants or BY-2 tobacco cell suspension cultures, which are transfected with recombinant Agrobacterium tumefaciens to induce transgene expression in the plant tissue. After 3–7 days of incubation, the recombinant protein can be extracted from the plant material. If required, the workflow from gene to purified protein can be achieved within 2 weeks. The versatility of the system allows the small‑scale expression screening of 20–40 constructs as well as the production of milligram amounts of proteins by the infiltration of larger quantities (up to 1 kg) of biomass. The N. benthamiana platform is currently being converted to a GMP-compliant manufacturing process.