Reference - Detail
|Author||Neumüller RA, Wirtz-Peitz F, Lee S, Kwon Y, Buckner M, Hoskins RA, Venken KJ, Bellen HJ, Mohr SE, Perrimon N.|
|Title||Stringent analysis of gene function and protein-protein interactions using fluorescently tagged genes.|
In Drosophila collections of green fluorescent protein (GFP) trap lines have been used to probe the endogenous expression patterns of trapped genes or the subcellular localization of their protein products. Here, we describe a method, based on nonoverlapping, highly specific, shRNA transgenes directed against GFP, that extends the utility of these collections to loss-of-function studies. Furthermore, we used a MiMIC transposon to generate GFP traps in Drosophila cell lines with distinct subcellular localization patterns, which will permit high-throughput screens using fluorescently tagged proteins. Finally, we show that fluorescent traps, paired with recombinant nanobodies and mass spectrometry, allow the study of endogenous protein complexes in Drosophila.
|MeSH||Animals Cell Line Cell Survival / genetics Drosophila / genetics* Drosophila / metabolism* Drosophila Proteins / genetics* Drosophila Proteins / metabolism* Embryo, Nonmammalian / metabolism Female Fluorescent Dyes* Gene Order Gene Silencing Green Fluorescent Proteins* / genetics Green Fluorescent Proteins* / metabolism Multiprotein Complexes / isolation & purification Multiprotein Complexes / metabolism Peptide Elongation Factors / genetics Peptide Elongation Factors / metabolism Protein Binding / physiology Protein Interaction Mapping / methods* RNA, Small Interfering / metabolism Recombinant Fusion Proteins / genetics Recombinant Fusion Proteins / metabolism Stem Cells / metabolism|
|WOS Category||GENETICS & HEREDITY|
|Drosophila||sgg-YFP (CPTI-002603 DGRC#115553)|