ArticleDevelopmental biology2017
Optimization of CRISPR/Cas9 genome editing for loss-of-function in the early chick embryo.
Article in Developmental biology, 2017. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 51 papers.
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Who cites it
51 citing papers in PubMed.
- Alternative Developmental Toxicity Screening Methods Using Chick Embryo-Based Models.Journal of applied toxicology : JAT · 2026Review
- A low-level Cdkn1c/p57EMBO reports · 2026Article
- The Neural Tube CURE: Engaging undergraduate students in a relevant developmental biology research course.Developmental biology · 2025Review
- Article
- CRISPR-Cas13d as a molecular tool to achieve targeted gene expression knockdown in chick embryos.Developmental biology · 2025Article
- CRISPR/Cas9-TargetedJournal of developmental biology · 2025Article
- An in vivo CRISPR screen in chick embryos reveals a role for MLLT3 in specification of neural cells from the caudal epiblast.Development (Cambridge, England) · 2025Article
- CRISPR/Cas9-based programmable genome editing in chickens: concepts, applications and regulatory issues.Frontiers in genome editing · 2025Review
- CRISPR-Cas13d as a molecular tool to achieve targeted gene expression knockdown in chick embryos.bioRxiv : the preprint server for biology · 2024Article
- Mitochondrial dynamics regulate cell morphology in the developing cochlea.Development (Cambridge, England) · 2024Article
- miR-203 secreted in extracellular vesicles mediates the communication between neural crest and placode cells required for trigeminal ganglia formation.PLoS biology · 2024Article
- CRISPR/Cas9-mediated knockout of the Vanin-1 gene in the Leghorn Male Hepatoma cell line and its effects on lipid metabolism.Animal bioscience · 2024Article
- Face processing in animal models: implications for autism spectrum disorder.Frontiers in neuroscience · 2024Review
- Article
- Temporal changes in plasma membrane lipid content induce endocytosis to regulate developmental epithelial-to-mesenchymal transition.Proceedings of the National Academy of Sciences of the United States of America · 2022Article
- Co-option of the piRNA pathway to regulate neural crest specification.Science advances · 2022Article
- Genome editing of avian species: implications for animal use and welfare.Laboratory animals · 2022Article
- Fourth Report on Chicken Genes and Chromosomes 2022.Cytogenetic and genome research · 2022Review
- In Ovo Gain- and Loss-of-Function Approaches to Study Gut Morphogenesis.Methods in molecular biology (Clifton, N.J.) · 2022Review
- Linking protein structural and functional change to mutation using amino acid networks.PloS one · 2022Article
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Authors and funding
4 authors.
Funding
Abstract
The advent of CRISPR/Cas9 has made genome editing possible in virtually any organism, including those not previously amenable to genetic manipulations. Here, we present an optimization of CRISPR/Cas9 for application to early avian embryos with improved efficiency via a three-fold strategy. First, we employed Cas9 protein flanked with two nuclear localization signal sequences for improved nuclear localization. Second, we used a modified guide RNA (gRNA) scaffold that obviates premature termination of transcription and unstable Cas9-gRNA interactions. Third, we used a chick-specific U6 promoter that yields 4-fold higher gRNA expression than the previously utilized human U6. For rapid screening of gRNAs for in vivo applications, we also generated a chicken fibroblast cell line that constitutively expresses Cas9. As proof of principle, we performed electroporation-based loss-of-function studies in the early chick embryo to knock out Pax7 and Sox10, key transcription factors with known functions in neural crest development. The results show that CRISPR/Cas9-mediated deletion causes loss of their respective proteins and transcripts, as well as predicted downstream targets. Taken together, the results reveal the utility of this optimized CRISPR/Cas9 method for targeted gene knockout in chicken embryos in a manner that is reproducible, robust and specific.
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