An expanded auxin-inducible degron toolkit for Caenorhabditis elegans. Issue 3 (20th January 2021)

• Record Type: Journal Article
• Authors: Ashley, Guinevere E; Duong, Tam; Levenson, Max T; Martinez, Michael A Q; Johnson, Londen C; Hibshman, Jonathan D; Saeger, Hannah N; Palmisano, Nicholas J; Doonan, Ryan; Martinez-Mendez, Raquel; Davidson, Brittany R; Zhang, Wan; Ragle, James Matthew; Medwig-Kinney, Taylor N; Sirota, Sydney S; Goldstein, Bob; Matus, David Q; Dickinson, Daniel J; Reiner, David J; Ward, Jordan D
• Editors: Greenstein, D
• Is Part Of: Genetics. Volume 217:Issue 3(2021)
• Publication Date: 2021-01-20
• Subjects: C. elegans -- AID system -- SapTrap -- self-excising cassette -- CRISPR/Cas9 -- Transport Inhibitor Response 1; Genetics -- Periodicals; 576.5
• DOI: 10.1093/genetics/iyab006
• Languages: English
• ISSNs: 0016-6731

Abstract

Abstract: The auxin-inducible degron (AID) system has emerged as a powerful tool to conditionally deplete proteins in a range of organisms and cell types. Here, we describe a toolkit to augment the use of the AID system in Caenorhabditis elegans . We have generated a set of single-copy, tissue-specific (germline, intestine, neuron, muscle, pharynx, hypodermis, seam cell, anchor cell) and pan-somatic TIR1 - expressing strains carrying a co-expressed blue fluorescent reporter to enable use of both red and green channels in experiments. These transgenes are inserted into commonly used, well-characterized genetic loci. We confirmed that our TIR1-expressing strains produce the expected depletion phenotype for several nuclear and cytoplasmic AID-tagged endogenous substrates. We have also constructed a set of plasmids for constructing repair templates to generate fluorescent protein::AID fusions through CRISPR/Cas9-mediated genome editing. These plasmids are compatible with commonly used genome editing approaches in the C. elegans community (Gibson or SapTrap assembly of plasmid repair templates or PCR-derived linear repair templates). Together these reagents will complement existing TIR1 strains and facilitate rapid and high-throughput fluorescent protein::AID tagging of genes. This battery of new TIR1-expressing strains and modular, efficient cloning vectors serves as a platform for straightforward assembly of CRISPR/Cas9 repair templates for conditional protein depletion.