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SYNCAS : efficient CRISPR/Cas9 gene-editing in difficult to transform arthropods

Sander De Rouck (UGent) , Antonio Mocchetti (UGent) , Wannes Dermauw (UGent) and Thomas Van Leeuwen (UGent)
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Abstract
The genome editing technique CRISPR/Cas9 has led to major advancements in many research fields and this state-of-the-art tool has proven its use in genetic studies for various arthropods. However, most transformation protocols rely on microinjection of CRISPR/Cas9 components into embryos, a method which is challenging for many species. Alternatively, injections can be performed on adult females, but transformation efficiencies can be very low as was shown for the two-spotted spider mite, Tetranychus urticae, a minute but important chelicerate pest on many crops. In this study, we explored different CRISPR/Cas9 formulations to optimize a maternal injection protocol for T. urticae. We observed a strong synergy between branched amphipathic peptide capsules and saponins, resulting in a significant increase of CRISPR/Cas9 knock-out efficiency, exceeding 20%. This CRISPR/ Cas9 formulation, termed SYNCAS, was used to knock-out different T. urticae genes - phytoene desaturase, CYP384A1 and Antennapedia - but also allowed to develop a co-CRISPR strategy and facilitated the generation of T. urticae knock-in mutants. In addition, SYNCAS was successfully applied to knock-out white and white-like genes in the western flower thrips, Frankliniella occidentalis. The SYNCAS method allows routine genome editing in these species and can be a game changer for genetic research in other hard to transform arthropods.
Keywords
Cas9 delivery, BAPC, Endosomal escape reagent, Hox genes, ABC-transporters, Chitin synthase, 2-SPOTTED SPIDER-MITE, TETRANYCHUS-URTICAE, GENOME, SAPONINS, RESISTANCE, DROSOPHILA, DELIVERY, IMPROVEMENTS, CHELICERATE, MUTATIONS

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MLA
De Rouck, Sander, et al. “SYNCAS : Efficient CRISPR/Cas9 Gene-Editing in Difficult to Transform Arthropods.” INSECT BIOCHEMISTRY AND MOLECULAR BIOLOGY, vol. 165, 2024, doi:10.1016/j.ibmb.2023.104068.
APA
De Rouck, S., Mocchetti, A., Dermauw, W., & Van Leeuwen, T. (2024). SYNCAS : efficient CRISPR/Cas9 gene-editing in difficult to transform arthropods. INSECT BIOCHEMISTRY AND MOLECULAR BIOLOGY, 165. https://doi.org/10.1016/j.ibmb.2023.104068
Chicago author-date
De Rouck, Sander, Antonio Mocchetti, Wannes Dermauw, and Thomas Van Leeuwen. 2024. “SYNCAS : Efficient CRISPR/Cas9 Gene-Editing in Difficult to Transform Arthropods.” INSECT BIOCHEMISTRY AND MOLECULAR BIOLOGY 165. https://doi.org/10.1016/j.ibmb.2023.104068.
Chicago author-date (all authors)
De Rouck, Sander, Antonio Mocchetti, Wannes Dermauw, and Thomas Van Leeuwen. 2024. “SYNCAS : Efficient CRISPR/Cas9 Gene-Editing in Difficult to Transform Arthropods.” INSECT BIOCHEMISTRY AND MOLECULAR BIOLOGY 165. doi:10.1016/j.ibmb.2023.104068.
Vancouver
1.
De Rouck S, Mocchetti A, Dermauw W, Van Leeuwen T. SYNCAS : efficient CRISPR/Cas9 gene-editing in difficult to transform arthropods. INSECT BIOCHEMISTRY AND MOLECULAR BIOLOGY. 2024;165.
IEEE
[1]
S. De Rouck, A. Mocchetti, W. Dermauw, and T. Van Leeuwen, “SYNCAS : efficient CRISPR/Cas9 gene-editing in difficult to transform arthropods,” INSECT BIOCHEMISTRY AND MOLECULAR BIOLOGY, vol. 165, 2024.
@article{01HMZWGRBXWD8ZR0YZNMGM5SC0,
  abstract     = {{The genome editing technique CRISPR/Cas9 has led to major advancements in many research fields and this state-of-the-art tool has proven its use in genetic studies for various arthropods. However, most transformation protocols rely on microinjection of CRISPR/Cas9 components into embryos, a method which is challenging for many species. Alternatively, injections can be performed on adult females, but transformation efficiencies can be very low as was shown for the two-spotted spider mite, Tetranychus urticae, a minute but important chelicerate pest on many crops. In this study, we explored different CRISPR/Cas9 formulations to optimize a maternal injection protocol for T. urticae. We observed a strong synergy between branched amphipathic peptide capsules and saponins, resulting in a significant increase of CRISPR/Cas9 knock-out efficiency, exceeding 20%. This CRISPR/ Cas9 formulation, termed SYNCAS, was used to knock-out different T. urticae genes - phytoene desaturase, CYP384A1 and Antennapedia - but also allowed to develop a co-CRISPR strategy and facilitated the generation of T. urticae knock-in mutants. In addition, SYNCAS was successfully applied to knock-out white and white-like genes in the western flower thrips, Frankliniella occidentalis. The SYNCAS method allows routine genome editing in these species and can be a game changer for genetic research in other hard to transform arthropods.}},
  articleno    = {{104068}},
  author       = {{De Rouck, Sander and Mocchetti, Antonio and Dermauw, Wannes and Van Leeuwen, Thomas}},
  issn         = {{0965-1748}},
  journal      = {{INSECT BIOCHEMISTRY AND MOLECULAR BIOLOGY}},
  keywords     = {{Cas9 delivery,BAPC,Endosomal escape reagent,Hox genes,ABC-transporters,Chitin synthase,2-SPOTTED SPIDER-MITE,TETRANYCHUS-URTICAE,GENOME,SAPONINS,RESISTANCE,DROSOPHILA,DELIVERY,IMPROVEMENTS,CHELICERATE,MUTATIONS}},
  language     = {{eng}},
  pages        = {{13}},
  title        = {{SYNCAS : efficient CRISPR/Cas9 gene-editing in difficult to transform arthropods}},
  url          = {{http://doi.org/10.1016/j.ibmb.2023.104068}},
  volume       = {{165}},
  year         = {{2024}},
}

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