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Setaria viridis, the wild ancestor of foxtail millet (Setaria italica), is an effective model plant for larger C4 crops because S. viridis has several desirable traits, such as short generation time, prolific seed production and a small genome size. These advantages are well suited for investigating molecular mechanisms in angiosperms, especially C4 crop species. Here, we report a procedure for isolating gametes and zygotes from S. viridis flowers. To isolate egg cells, ovaries were harvested from unpollinated mature flowers and cut transversel