Agrobacterium-mediatedCuscuta campestristransformation as a tool for understanding plant-plant interactions

Abstract

SummaryCuscuta campestris, a stem parasitic plant, has served as a valuable model plant for the exploration of plant-plant interactions and molecular trafficking. However, a major barrier toC. campestrisresearch is that a method to generate stable transgenic plants has not yet been developed. Here, we describe the development of aCuscutatransformation protocol using various reporter genes (GFP, GUS, or RUBY) and morphogenic genes (CcWUS2andCcGRF/GIF), ultimately leading to a robust protocol forAgrobacterium-mediatedC. campestristransformation. The stably transformed and regenerated RUBYC. campestrisplants produced haustoria, the signature organ of parasitic plants, and these were functional in forming host attachments. The locations of T-DNA integration in the parasite genome were confirmed through TAIL-PCR. TransformedC. campestrisalso produced flowers and transgenic seeds exhibiting betalain pigment, providing proof of germline transmission of the RUBY transgene. Furthermore, the RUBY reporter is not only a useful selectable marker for theAgrobacterium-mediated transformation, but also provides insight into the movement of molecules fromC. campestristo the host during parasitism. Thus, the protocol for transformation ofC. campestrisreported here overcomes a major obstacle toCuscutaresearch and opens new possibilities for studying parasitic plants and their interactions with hosts.