Developing Tools for Studying Roots in Setaria

Setaria viridis is a spectacularly successful invasive grass that grows quickly to maturity even in low water, soil fertility or space. It is a model plant for switchgrass, maize, and foxtail millet, its domesticated relative.

  We are studying Setaria as a model for monocot root development (see poster by Sebastian, J.) and response to stress. We have previously shown that specific cell types are important for responses to salt stress in Arabidopsis roots. In order to facilitate identification and purification of root cell types in Setaria for developmental and transcriptomic analysis, we will make cell type-specific promoters linked to reporters or epitope tags. Expression will be studied both by fluorescence microscopy using  GFP and YFP reporters, and by in situ imaging of soil-grown plants in the GLO_ROOTS system using luciferase reporters (see presentation in Plant Phenotyping Workshop by Ruben Rellan.) Cell type-specific promoters fused to epitope tags for immunoprecipitation of ribosomal particles will enable us to follow abundance of translated mRNAs during drought or salt  stress.

 We have modified the pANIC vector by replacing the Gateway recombination sites with a Type IIs restriction enzyme for use in Golden Gate cloning. This facilitates the rapid and inexpensive generation of multi-component transgenes in one step and yields “scarless” recombinant plasmids. We are currently cloning promoters from Setaria and Arabidopsis into this pGolden-Gate pANIC vector and the Arabidopsis binary vector pGoldenGate-MCY2 for testing reciprocal expression in both plants.