interactions Protocols

Category: RNAi Technology Protocols: RNAi C. elegans Protocols

Protocol for RNAi screens in C. elegans in a 96-well liquid format and their application to the systematic identification of genetic interactions. The procedure allows thousands of RNAi feeding experiments to be performed per investigator per day. - [Read RNAi Screens in C. elegans Protocol]

Category: Reporter Gene Assays: Luciferase Assay Protocols

Protocol describes a split luciferase complementation assay used to study the interaction of proteins in cells. In the split protein strategy, a single reporter protein/enzyme (firefly luciferase [Fluc]) is cleaved into amino-terminal and carboxy-terminal halves; each half is fused to one of two interacting proteins, X & Y. Physical interactions between the two proteins reconstitute the functional reporter protein, leading to enzymatic activities that can be measured by in vitro or in vivo assay - [Read Split Luciferase Complementation Assay for Studying Interaction of Proteins X and Y in Cells]

Category: Cell Biology and Cell Culture: In Vitro Reconstitution Protocols

Small ubiquitin-like modifier (SUMO) is a small molecule, but has a variety of regulatory functions in cells. SUMO modification is involved in transcriptional regulation, subcellular localization, and protein-protein interactions. SUMO conjugation requires sequential E1-dependent activation, E2-dependent conjugation, and E3-dependent ligation steps. Protocol includes: In vivo and in vitro SUMOylation assay and deSUMOylation assay. - [Read Sumoylation and Desumoylation Assays for a Chromatin-Remodelling Complex In Vivo and In Vitro]

Category: Immunology: Immunoprecipitation Protocols: Protein Immunoprecipitation Protocols

Protocol describes the use of FLAG-epitope-tagged proteins for both small-scale analysis and large-scale coimmunoprecipitation of interacting proteins. When examining protein interactions, it is sometimes possible to immunoprecipitate an endogenous protein X directly, without using an epitope tag, if antibodies are available. The advantage of examining interactions of endogenously expressed proteins is that these are likely to be physiological and less likely to be an artifact of overexpression. - [Read Using FLAG Epitope-Tagged Proteins for Coimmunoprecipitation of Interacting Proteins Protocol]

Category: Molecular Model Organisms: Yeast Two-Hybrid Screen Protocol

Yeast Two-Hybrid: State of the Art. Paper on Protein-protein interactions and the two-hybrid protocol. Wim Van Criekinge et al., BPO. - [Read Yeast Two-Hybrid: State of the Art]