Genetic screening of mutants and identification of their causative genes are effective strategies for understanding plant signaling pathways. Here, we present Lumi-Map, a method that combines real-time luciferase bioluminescence monitoring with MutMap analysis to identify single nucleotide polymorphisms (SNPs) responsible for the mutant phenotypes. Real-time monitoring of reporter gene expression enables high-throughput screen of mutants with altered transcriptional responses. Once candidate mutants are isolated, a whole-genome sequencing-based method MutMap allows for rapid identification of the causative SNPs. Using reporter lines driven by defense-related gene promoters, Lumi-Map successfully identified genes involved in pattern-triggered immunity, including a novel receptor gene for pathogen-associated molecular patterns. By selecting appropriate luciferase reporter plants, Lumi-Map is expected to serve as a versatile platform for dissecting diverse signal transduction pathways in plants.