Forward Chemical Screening of Small RNA Pathways

RNA silencing is a mechanism of gene expression regulation mediated by short noncoding RNAs called small RNAs. Small RNAs can suppress gene expression transcriptionally or posttranscriptionally by base pairing to their targets. In plants, they are involved in a diverse range of pathways and processes. Forward genetic screens have led to the identification of many components in small RNA biogenesis and functional pathways. However, it cannot identify essential or functionally redundant genes. Forward chemical screens can overcome these limitations by targeting multiple redundant proteins within a family and by giving the user temporal control of the application of the chemical. Here, we describe a method to quickly screen chemicals that perturb small RNA pathways using Arabidopsis thaliana reporter lines in a 96-well format.