Agrobacterium Protocols

Joyce Van Eck, Kerry Swartwood, Joyce Van Eck Ph.D., Eck, Joyce Van, Swartwood, Kerry

Interest in plant model systems for genetic, biological, and functional genomics studies stems from advantages they provide in terms of fast generation time, small stature, and simple growth requirements. A model species would be especially advantageous for the studies of C4 photosynthetic grasses, which currently present practical challenges. These include long seed-to-seed generation times, and because of their large size at maturity they require large growing areas. One potential model of interest for C4 photosynthetic grasses is Setaria viridis. It has all the desirable aforementioned attributes for a model; however, for it to be adopted as a model for functional genomics studies, gene transfer methodology is also needed. In this chapter, we describe methods for Agrobacterium tumefaciens-mediated transformation of seed-derived callus. Vectors used for gene constructs contained the hygromycin phosphotransferase gene as a selectable marker. The transition of transgenic lines to soil was straightforward; plants started to flower in 1-3 weeks, with seeds ready to harvest approximately 5 weeks later.