The Analysis of Protein-Protein Interactions in Plants by Bimolecular Fluorescence Complementation

The Analysis of Protein-Protein Interactions in Plants by Bimolecular Fluorescence Complementation The BiFC Principle BiFC is based upon tethering split YFP or other GFP variants to form a functional fluorophore. The association of the split YFP/GFP/cyan fluorescent protein (CFP) molecule does not occur spontaneously and requires interaction between proteins or peptides that are fused to each of the fluorophore fragments ( Fig. 1 ). Upon interaction of these fused proteins/peptides, the split fluorophore fragments can interact to form a fluorescent protein that has the same spectral properties as the unsplit YFP (or other GFP variants; Figs. 1 and 2 ). If the proteins that are fused to the split fluorophore fragments do not interact, reconstitution of the YFP/GFP/CFP usually does not take place and no fluorescence is detected. View larger version (31K): In this window In a new window As a PowerPoint slide Figure 1. BiFC can be used to determine subcellular localization of protein complexes. A, Under physiological conditions reconstitution of a fluorescent YFP molecule can only take place following interaction between proteins or peptides that are fused to YN and YC fragments. B, A BiFC assay showing that complexes of AtROP6 and the ROP-interacting coiled-coil scaffold protein ICR1 are localized in the plasma membrane of N. benthamiana http://www.deepdyve.com/assets/images/DeepDyve-Logo-lg.png

The Analysis of Protein-Protein Interactions in Plants by Bimolecular Fluorescence Complementation

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Publisher
American Society of Plant Biologist
Copyright
Copyright © 2015 by the American Society of Plant Biologists
ISSN
1532-2548
eISSN
0032-0889
D.O.I.
10.1104/pp.107.107284
Publisher site
See Article on Publisher Site

Abstract

The BiFC Principle BiFC is based upon tethering split YFP or other GFP variants to form a functional fluorophore. The association of the split YFP/GFP/cyan fluorescent protein (CFP) molecule does not occur spontaneously and requires interaction between proteins or peptides that are fused to each of the fluorophore fragments ( Fig. 1 ). Upon interaction of these fused proteins/peptides, the split fluorophore fragments can interact to form a fluorescent protein that has the same spectral properties as the unsplit YFP (or other GFP variants; Figs. 1 and 2 ). If the proteins that are fused to the split fluorophore fragments do not interact, reconstitution of the YFP/GFP/CFP usually does not take place and no fluorescence is detected. View larger version (31K): In this window In a new window As a PowerPoint slide Figure 1. BiFC can be used to determine subcellular localization of protein complexes. A, Under physiological conditions reconstitution of a fluorescent YFP molecule can only take place following interaction between proteins or peptides that are fused to YN and YC fragments. B, A BiFC assay showing that complexes of AtROP6 and the ROP-interacting coiled-coil scaffold protein ICR1 are localized in the plasma membrane of N. benthamiana

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