Establishment of a CRISPR/Cas9-based strategy for inducible protein dimerization

Zielich, Jeffrey; Mangal, Sriyash; Zanin, Esther; Lambie, Eric J.

doi:10.17912/w2208r

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2020

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“…Phenotype analysis and microscopy for worm experiments. Rapamycin and DMSO microinjections were performed as described 60 . Once adults were injected with different reagents (5% DMSO and 100 µM, 200 µM, 300 µM rapamycin or without injection), they were imaged using a Leica TCS SP5 II confocal microscope at 0, 1, 2, 3, 4, 5, 6, 12, 18, 24, and 30 h after injection.…”

Section: Data Availabilitymentioning

confidence: 99%

Tunable light and drug induced depletion of target proteins

et al. 2020

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Biological processes in development and disease are controlled by the abundance, localization and modification of cellular proteins. We have developed versatile tools based on recombinant E3 ubiquitin ligases that are controlled by light or drug induced heterodimerization for nanobody or DARPin targeted depletion of endogenous proteins in cells and organisms. We use this rapid, tunable and reversible protein depletion for functional studies of essential proteins like PCNA in DNA repair and to investigate the role of CED-3 in apoptosis during Caenorhabditis elegans development. These independent tools can be combined for spatial and temporal depletion of different sets of proteins, can help to distinguish immediate cellular responses from long-term adaptation effects and can facilitate the exploration of complex networks.

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Section: Data Availabilitymentioning

confidence: 99%