Staff Research Associate Univeristy of California, Berkelely Oakland, California
Body of Abstract: Setaria viridis, commonly known as green foxtail, is a C4 grass species and a close relative of major cereal crops, including maize and sorghum. Its small diploid genome, short stature, rapid life cycle, and availability of quality genome sequences make it an ideal model plant to study C4 crops. However, absence of efficient genome engineering and editing strategies for Setaria viridis limits its utility as a model plant. In this study, we used two different Agrobacterium-mediated transformation strategies. One of the strategies used dry seed-derived callus of ME034 as a target tissue for transformation. Using a pANIC 10A construct, which contains the plant selectable marker gene hygromycin B phosphotransferase and a visual marker gene of red fluorescent protein, we achieved a transformation efficiency of 31.0%. For genome editing, CRISPR/Cas9 together with a tRNA-gRNA system was applied to edit the Seteria endogenous gene, phytoene desaturase. This strategy led to an efficient editing and knockout rate of 67.9% and 46.4%, respectively. We also employed a transformation strategy that used A10.1 dry seed explants as a target tissue for Agrobacterium-based transformation using a construct containing maize morphogenic genes Baby boom and Wuschel2. Previous studies have shown that using these developmental genes improves transformation and editing efficiency in different cereal crops. This dry seed-based strategy avoids the necessity of developing callus to use as a target tissue for transformation. It also shortens the time to generate transgenic T0 plants by four-six weeks, compared to callus-based transformation. Using this strategy, an average transformation efficiency of 11.0 % was achieved. Both the callus and dry seed-based strategies are valuable tools to broaden the utility of Setaria viridis as a model C4 crop.