ArticleG3 (Bethesda, Md.)2014
A versatile two-step CRISPR- and RMCE-based strategy for efficient genome engineering in Drosophila.
Article in G3 (Bethesda, Md.), 2014. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 72 papers.
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72 citing papers in PubMed.
- Genetic analyses enabled by the fourth chromosome resource project reveal unexpected mutant phenotypes and suggest new disease models.G3 (Bethesda, Md.) · 2026Article
- The mRNA export factor UAP56 is required for dendrite and synapse pruning via actin regulation in Drosophila.Journal of cell science · 2026Article
- Decoding the molecular logic of rapidly evolving ZAD zinc finger proteins inScience advances · 2026Article
- Trans-Splicing Enhances Processivity of RNA Polymerase II and Suppresses Polyadenylation Signals.Doklady. Biochemistry and biophysics · 2026Article
- Functional profiling and visualization of the sphingolipid metabolic network in vivo.EMBO reports · 2025Article
- dFlpTag, a RMCE-based tool for simultaneous endogenous protein tagging and cell labeling in Drosophila.Scientific reports · 2025Article
- Catalytic-dependent and independent functions of the histone acetyltransferase CBP promote pioneer-factor-mediated zygotic genome activation.Molecular cell · 2025Article
- Titin-dependent biomechanical feedback tailors sarcomeres to specialized muscle functions in insects.Science advances · 2025Article
- Investigation of the Functional Role of the Conserved Sequence at the 5'-End of the Fourth Intron of the mod(mdg4) Gene in Trans-Splicing in Drosophila melanogaster.Doklady. Biochemistry and biophysics · 2025Article
- Contributions of the Dachsous intracellular domain to Dachsous-Fat signaling.Development (Cambridge, England) · 2024Article
- The Hox protein Antennapedia orchestratesScience advances · 2024Article
- Bruno 1/CELF regulates splicing and cytoskeleton dynamics to ensure correct sarcomere assembly in Drosophila flight muscles.PLoS biology · 2024Article
- Enhancer-promoter interactions can form independently of genomic distance and be functional across TAD boundaries.Nucleic acids research · 2024Article
- Unveiling the Genetic Symphony: Harnessing CRISPR-Cas Genome Editing for Effective Insect Pest Management.Plants (Basel, Switzerland) · 2023Review
- CRISPR/Cas9-mediated knock-in inGene and genome editing · 2023Article
- Systematic analysis of RhoGEF/GAP localizations uncovers regulators of mechanosensing and junction formation during epithelial cell division.Current biology : CB · 2023Article
- Parkinsonism mutations in DNAJC6 cause lipid defects and neurodegeneration that are rescued by Synj1.NPJ Parkinson's disease · 2023Article
- TheeLife · 2022Article
- Article
- Speciation and adaptation research meets genome editing.Philosophical transactions of the Royal Society of London. Series B, Biological sciences · 2022Review
12 more citing papers are in PubMed but not listed here.
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3 authors.
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No grant is acknowledged in the PubMed record.
Abstract
The development of clustered, regularly interspaced, short palindromic repeats (CRISPR)/CRISPR-associated (Cas) technologies promises a quantum leap in genome engineering of model organisms. However, CRISPR-mediated gene targeting reports in Drosophila melanogaster are still restricted to a few genes, use variable experimental conditions, and vary in efficiency, questioning the universal applicability of the method. Here, we developed an efficient two-step strategy to flexibly engineer the fly genome by combining CRISPR with recombinase-mediated cassette exchange (RMCE). In the first step, two sgRNAs, whose activity had been tested in cell culture, were co-injected together with a donor plasmid into transgenic Act5C-Cas9, Ligase4 mutant embryos and the homologous integration events were identified by eye fluorescence. In the second step, the eye marker was replaced with DNA sequences of choice using RMCE enabling flexible gene modification. We applied this strategy to engineer four different locations in the genome, including a gene on the fourth chromosome, at comparably high efficiencies. Our data suggest that any fly laboratory can engineer their favorite gene for a broad range of applications within approximately 3 months.
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