Compartmentalized Terpenoid Production in Plants Using Agrobacterium-Mediated Transient Expression

As the field of plant synthetic biology continues to grow, Agrobacterium-mediated transient expression has become an essential method to rapidly test pathway candidate genes in a combinatorial fashion. This is especially important when elucidating and engineering more complex pathways to produce commercially relevant chemicals like many terpenoids, a widely diverse class of natural products of often industrial relevance. Agrobacterium-mediated transient expression has facilitated multiplex expression of recombinant and modified enzymes, including synthetic biology approaches to compartmentalize the biosynthesis of terpenoids subcellularly. Here, we describe methods on how to deploy Agrobacterium-mediated transient expression in Nicotiana benthamiana to rapidly develop terpenoid pathways and compartmentalize terpenoid biosynthesis within plastids, the cytosol, or at the surface of lipid droplets.