title: Quantitative analysis of protein-protein interactions by split firefly luciferase complementation in plant protoplasts.
abstract: This unit describes the split firefly luciferase complementation (SFLC) assay, a high-throughput quantitative method that can be used to investigate protein-protein interactions (PPIs) in plant mesophyll protoplasts. In SFLC, the two proteins to be tested for interaction are expressed as chimeric proteins, each fused to a different half of firefly luciferase. If the proteins interact, a functional luciferase can be transitorily reconstituted, and is detected using the cell-permeable substrate D-luciferin. An advantage of the SFLC assay is that dynamic changes in PPIs in a cell can be detected in a near real-time manner. Another advantage is the unusually high DNA co-transfection and protein expression efficiencies that can be achieved in plant protoplasts, thereby enhancing the throughput of the method.
authors.affiliation: Department of Molecular Biology, Massachusetts General Hospital, Boston, Massachusetts.
authors.forename: Jian-Feng
authors.initials: JF
authors.lastname: Li
authors.affiliation: Department of Molecular Biology, Massachusetts General Hospital, Boston, Massachusetts.
authors.forename: Dandan
authors.initials: D
authors.lastname: Zhang
mesh_terms: D015870:Gene Expression; D005816:Genetic Complementation Test / Q000379:methods*; D049409:Luciferases, Firefly / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D059828:Plant Cells / Q000378:metabolism*; D010940:Plant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011523:Protoplasts / Q000166:cytology / Q000378:metabolism*; D014162:Transfection / Q000379:methods
chemical_list: D010940:Plant Proteins; D049409:Luciferases, Firefly
keywords: plant protoplasts; protein-protein interaction; split firefly luciferase complementation