65 Friedrich Fauser and Martin Jonikas (eds.), Plant Chemical Genomics: Methods and Protocols, Methods in Molecular Biology, vol. 1795, https://doi.org/10.1007/978-1-4939-7874-8_6, © Springer Science+Business Media, LLC, part of Springer Nature 2018 Chapter 6 Identifcation of Chemical Inducers of the  Phosphate-Starvation Signaling Pathway in A. thaliana Using Chemical Genetics Clémence Bonnot, Laurent Nussaume, and Thierry Desnos Abstract In spite of its importance for agriculture and 30 years of genetic studies, the phosphate-starvation signaling pathway, that allows plants to detect, respond, and adapt to changes in the phosphate concentration of the rhizosphere, remains poorly known. Chemical genetics has been increasingly and successfully used as a complementary approach to genetics for the dissection of signaling pathways in diverse organisms. Screens can be designed to identify chemicals interfering specifcally with a pathway of interest. We designed a screen that led to the discovery of the frst chemical capable to induce specifcally the phosphate-starvation signaling pathway in Arabidopsis thaliana. This procedure, described here, can be adapted for the discovery of inducers or repressors of other pathways. Key words Chemical genetics screen, Reporter gene, Phosphate-starvation signaling pathway, Structure–activity relationship, Metabolization, Degradation, Mass-spectrometry, Arabidopsis thaliana 1 Introduction Chemical genetics is a method of investigation of biological processes consisting in identifying and using small molecules (<500 Da), of synthetic or natural origin, to disturb a biological activity in a specifc manner. Similarly to mutations in classical genetics, the chemicals used in chemical genetics induce gain or loss or alter partially the function of their target (protein, RNA, etc.) [1–3]. The analysis of the phenotypes generated and the identifcation of the target can be used for the discovery of new genes and functions [4–7]. In plant sciences, chemical genetics was historically used for the discovery and improvement of agrochemicals such as herbicides, and for the analysis of their mode of actions [8]. However, over the last 20 years, chemical genetics has been increasingly and successfully applied to basic research as a complementary approach to genetics [7, 9, 10]. Small molecules were instrumental in the discovery of novel molecular components of several biological pathways