1. A library of MiMICs allows tagging of genes and reversible, spatial and temporal knockdown of proteins in Drosophila.
- Author
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Nagarkar-Jaiswal S, Lee PT, Campbell ME, Chen K, Anguiano-Zarate S, Gutierrez MC, Busby T, Lin WW, He Y, Schulze KL, Booth BW, Evans-Holm M, Venken KJ, Levis RW, Spradling AC, Hoskins RA, and Bellen HJ
- Subjects
- Animals, Animals, Genetically Modified, Blotting, Western, Brain metabolism, Drosophila Proteins metabolism, Drosophila melanogaster metabolism, Drosophila melanogaster physiology, Gene Expression, Green Fluorescent Proteins genetics, Green Fluorescent Proteins metabolism, Larva genetics, Larva metabolism, Learning physiology, Microscopy, Confocal, Time Factors, Tumor Suppressor Proteins genetics, Tumor Suppressor Proteins metabolism, alpha Catenin genetics, alpha Catenin metabolism, DNA Transposable Elements genetics, Drosophila Proteins genetics, Drosophila melanogaster genetics, Gene Library, Mutagenesis, Insertional, RNA Interference
- Abstract
Here, we document a collection of ∼7434 MiMIC (Minos Mediated Integration Cassette) insertions of which 2854 are inserted in coding introns. They allowed us to create a library of 400 GFP-tagged genes. We show that 72% of internally tagged proteins are functional, and that more than 90% can be imaged in unfixed tissues. Moreover, the tagged mRNAs can be knocked down by RNAi against GFP (iGFPi), and the tagged proteins can be efficiently knocked down by deGradFP technology. The phenotypes associated with RNA and protein knockdown typically correspond to severe loss of function or null mutant phenotypes. Finally, we demonstrate reversible, spatial, and temporal knockdown of tagged proteins in larvae and adult flies. This new strategy and collection of strains allows unprecedented in vivo manipulations in flies for many genes. These strategies will likely extend to vertebrates.
- Published
- 2015
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