Abstract
In multicellular organisms, establishing the full body plane involves cell-cell signaling where protein associations are important for the diverse cellular functions within the cells. For the study of protein–protein interactions (PPI), bimolecular fluorescence complementation (BiFC) and luciferase complementation assays (LCA) have proven to be reliable tools that can be used to confirm the physical association of two proteins in a semi-in vivo environment. This chapter provides a detailed description of these two techniques using Nicotiana benthamiana as a semi-in vivo transient expression system. As an example, we will use the interaction of the two well-described transcription factors SHORT-ROOT (SHR) and SCARECROW (SCR), which are known as regulators of asymmetric cell division and stem cell specification in the root meristem of the model plant Arabidopsis thaliana. While the BiFC assay provides subcellular information by displaying a fluorescence signal, nuclear in this case, resulting from the reconstituted fluorophore, the LCA generates a quantitative readout of the SCR–SHR interaction. The combination of both assays provides information on the localization and strength of the PPI.
Original language | English (US) |
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Title of host publication | Methods in Molecular Biology |
Publisher | Humana Press Inc. |
Pages | 121-131 |
Number of pages | 11 |
DOIs | |
State | Published - 2023 |
Publication series
Name | Methods in Molecular Biology |
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Volume | 2690 |
ISSN (Print) | 1064-3745 |
ISSN (Electronic) | 1940-6029 |
Bibliographical note
Funding Information:This study was supported by KAUST baseline research funding to I.B. The authors declare no competing financial interests.
Publisher Copyright:
© 2023, The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.
Keywords
- BiFC
- Fluorescence
- LCA
- Luciferase
- Luminescence
- PPIs
- Tobacco
- YFP
ASJC Scopus subject areas
- Molecular Biology
- Genetics