The article describes the transient gene expression system using Arabidopsis mesophyll protoplasts, a versatile tool for conducting cell-based experiments in molecular, cellular, biochemical, genetic, genomic, and proteomic approaches. The method involves protoplast isolation, PEG–calcium transfection of plasmid DNA, and protoplast culture. This system offers physiological responses and high-throughput capability, enabling cost-effective explorations and hypothesis-driven tests. The protocol includes detailed steps for protoplast isolation, DNA transfection, and culture, with troubleshooting advice. The TEAMP system has been widely used to study various signaling pathways and cellular machineries in plant physiology, immunity, growth, and development. Advantages include uniform and abundant protoplasts, rapid response to diverse signals, and the ability to study processes beyond leaf functions. Limitations include the need for specialized growth conditions for long-term responses and the requirement for high transfection efficiency. The article also provides a list of useful reporter constructs and discusses the application of the TEAMP assay in functional genomics research.The article describes the transient gene expression system using Arabidopsis mesophyll protoplasts, a versatile tool for conducting cell-based experiments in molecular, cellular, biochemical, genetic, genomic, and proteomic approaches. The method involves protoplast isolation, PEG–calcium transfection of plasmid DNA, and protoplast culture. This system offers physiological responses and high-throughput capability, enabling cost-effective explorations and hypothesis-driven tests. The protocol includes detailed steps for protoplast isolation, DNA transfection, and culture, with troubleshooting advice. The TEAMP system has been widely used to study various signaling pathways and cellular machineries in plant physiology, immunity, growth, and development. Advantages include uniform and abundant protoplasts, rapid response to diverse signals, and the ability to study processes beyond leaf functions. Limitations include the need for specialized growth conditions for long-term responses and the requirement for high transfection efficiency. The article also provides a list of useful reporter constructs and discusses the application of the TEAMP assay in functional genomics research.