4.4 Article

An expanded auxin-inducible degron toolkit for Caenorhabditis elegans

Journal

GENETICS
Volume 217, Issue 3, Pages -

Publisher

GENETICS SOCIETY AMERICA
DOI: 10.1093/genetics/iyab006

Keywords

C. elegans; AID system; SapTrap; self-excising cassette; CRISPR/Cas9; Transport Inhibitor Response 1

Funding

  1. NIH/NIGMS [R00 GM107345, R01 GM138701]
  2. National Science Foundation (NSF) Division of Molecular and Cellular Biosciences (CAREER award) [1942922]
  3. NIH NIGMS [R01 GM121597]
  4. Damon Runyon Cancer Research Foundation [DRR-47-17]
  5. NIGMS [R01 GM121597-02S2]
  6. American Cancer Society [132969-PF-18-22601-CSM]
  7. NICHD [F31 HD100091]
  8. Cancer Prevention and Research Institute of Texas Grant [RR170054]
  9. NIH Office of Research Infrastructure Programs [P40 OD010440]
  10. [F32 GM131577]
  11. [R35 GM134838]
  12. [R00-GM115964]
  13. [R01 GM121625]
  14. Direct For Biological Sciences [1942922] Funding Source: National Science Foundation
  15. Div Of Molecular and Cellular Bioscience [1942922] Funding Source: National Science Foundation

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This study presents a toolkit for enhancing the application of the AID system in Caenorhabditis elegans, including tissue-specific and pan-somatic TIR1-expressing strains, as well as plasmids for constructing repair templates for generating fluorescent protein::AID fusions. These reagents will complement existing TIR1 strains and facilitate rapid and high-throughput tagging of genes with fluorescent protein::AID.
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.

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