Localizing protein-protein interactions by bimolecular fluorescence complementation in planta.
The application of novel assays for basic cell research is tightly linked to the development of easy-to-use and versatile tools and protocols for implementing such technologies for a wide range of applications and model species. The bimolecular fluorescence complementation (BiFC) assay is one such novel method for which tools and protocols for its application in plant cell research are still being developed. BiFC is a powerful tool which enables not only detection, but also visualization and subcellular localization of protein-protein interactions in living cells. Here we describe the application of BiFC in plant cells while focusing on the use of our versatile set of vectors which were specifically designed to facilitate the transformation, expression and imaging of protein-protein interactions in various plant species. We discuss the considerations of using our system in various plant model systems, the use of single versus multiple expression cassettes, the application of our vectors using various transformation methods and the use of internal fluorescent markers which can assist in signal localization and easy data acquisition in living cells.
Citovsky
Vitaly
V
Department of Biochemistry and Cell Biology, State University of New York, Stony Brook, NY 11794-5215, USA.
Gafni
Yedidya
Y
Tzfira
Tzvi
T
D001681:Biological Assay / Q000379:methods; D015415:Biomarkers / Q000032:analysis; D005456:Fluorescent Dyes / Q000032:analysis / Q000378:metabolism; D005822:Genetic Vectors; D008164:Luminescent Proteins / Q000032:analysis* / Q000235:genetics / Q000378:metabolism; D008856:Microscopy, Fluorescence / Q000379:methods*; D016254:Mutagenesis, Insertional; D010944:Plants / Q000235:genetics / Q000378:metabolism*; D010957:Plasmids; D011485:Protein Binding; D025941:Protein Interaction Mapping / Q000379:methods*; D011523:Protoplasts / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000032:analysis / Q000378:metabolism / Q000528:radiation effects; D014162:Transfection; D014170:Transformation, Genetic
D015415:Biomarkers; D005456:Fluorescent Dyes; D008164:Luminescent Proteins; D011993:Recombinant Fusion Proteins