Abstract
The identification of chemical compounds that affect intracellular processes has greatly contributed to the understanding of developmental regulation in plants. In this protocol, we describe a method for identifying chemical compounds that affect cold-regulated gene expression in Arabidopsis thaliana. Specifically, we generated Arabidopsis plants harboring a COLD-REGULATED 15A (COR15A) promoter::luciferase (COR15Apro::LUC) construct and grew them in a submerged liquid culture. Using a single true leaf excised from COR15Apro::LUC plants and 96-well culture plates, we performed high-throughput screening of chemical compounds that inhibit cold-induction of COR15Apro::LUC. Luciferase activity was detected using a microplate reader and a chemiluminescence imaging device. This protocol can be easily used for the identification of chemical compounds that regulate other processes, being versatile with respect to equipment. Key features • High-throughput screening of chemical compounds that affect cold-regulated gene expression is possible using a single leaf excised from Arabidopsis grown in a submerged culture. • Screening is based on luciferase activity derived from an excised single leaf. • Direct measurement of luciferase activity is possible using a microplate reader and a chemiluminescence imaging device. • This protocol can be easily used for the identification of chemical compounds that regulate other processes.