ArticleDevelopmental biology2017
Evaluation and rational design of guide RNAs for efficient CRISPR/Cas9-mediated mutagenesis in Ciona.
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 46 papers.
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Who cites it
46 citing papers in PubMed, 78 citations in OpenAlex.
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- Fine-tuning neuron number:bioRxiv : the preprint server for biology · 2026Article
- Neural signaling contributes to heart formation and growth in the invertebrate chordate, Ciona robusta.PLoS biology · 2026Article
- Insights into cephalochordate genome and gene evolution from the early-diverging amphioxusProceedings of the National Academy of Sciences of the United States of America · 2026Article
- A protocadherin mediates oral placode morphogenesis in the tunicate Ciona.Biology open · 2026Article
- An NKX2-5 homolog is required downstream of BMP signaling to pattern the sensory-adhesive organ of a tunicate larva.bioRxiv : the preprint server for biology · 2025Article
- Regulation of motor neuron differentiation in thebioRxiv : the preprint server for biology · 2025Article
- Cell cycle-driven transcriptome maturation confers multilineage competence to cardiopharyngeal progenitors.The EMBO journal · 2025Article
- Development of cell labeling and gene editing tools in urochordateMarine life science & technology · 2025Article
- RNA-seq analysis of wild-type and mutated TBPL1 gene in breast cancer cells lines through CRISPR/Cas9 approach reveals novel molecular signatures.Scientific reports · 2025Article
- Neuron-specific Agrin splicing by Nova RNA-binding proteins regulates conserved neuromuscular junction development in chordates.PLoS biology · 2025Article
- A gene regulatory network for specification and morphogenesis of a Mauthner Cell homolog in non-vertebrate chordates.Developmental biology · 2025Article
- The Evolution of Nucleic Acid-Based Diagnosis Methods from the (pre-)CRISPR to CRISPR era and the Associated Machine/Deep Learning Approaches in Relevant RNA Design.Methods in molecular biology (Clifton, N.J.) · 2025Article
- Structure-optimized sgRNA selection with PlatinumCRISPr for efficient Cas9 generation of knockouts.Genome research · 2024Article
- CRISPR/Cas9 Protocols for Disrupting Gene Function in the Non-vertebrate Chordate Ciona.Integrative and comparative biology · 2024Review
- A change in cis-regulatory logic underlying obligate versus facultative muscle multinucleation in chordates.Development (Cambridge, England) · 2024Article
- A conserved RNA switch for acetylcholine receptor clustering at neuromuscular junctions in chordates.bioRxiv : the preprint server for biology · 2024Article
- A change inbioRxiv : the preprint server for biology · 2024Article
- Specification of distinct cell types in a sensory-adhesive organ important for metamorphosis in tunicate larvae.PLoS biology · 2024Article
- Clustered Regularly Interspaced Short Palindromic Repeat/CRISPR-Associated Protein and Its Utility All at Sea: Status, Challenges, and Prospects.Microorganisms · 2024Review
Corrections and comments
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Authors and funding
5 authors at 2 institutions in 1 country.
Funding
Abstract
The CRISPR/Cas9 system has emerged as an important tool for various genome engineering applications. A current obstacle to high throughput applications of CRISPR/Cas9 is the imprecise prediction of highly active single guide RNAs (sgRNAs). We previously implemented the CRISPR/Cas9 system to induce tissue-specific mutations in the tunicate Ciona. In the present study, we designed and tested 83 single guide RNA (sgRNA) vectors targeting 23 genes expressed in the cardiopharyngeal progenitors and surrounding tissues of Ciona embryo. Using high-throughput sequencing of mutagenized alleles, we identified guide sequences that correlate with sgRNA mutagenesis activity and used this information for the rational design of all possible sgRNAs targeting the Ciona transcriptome. We also describe a one-step cloning-free protocol for the assembly of sgRNA expression cassettes. These cassettes can be directly electroporated as unpurified PCR products into Ciona embryos for sgRNA expression in vivo, resulting in high frequency of CRISPR/Cas9-mediated mutagenesis in somatic cells of electroporated embryos. We found a strong correlation between the frequency of an Ebf loss-of-function phenotype and the mutagenesis efficacies of individual Ebf-targeting sgRNAs tested using this method. We anticipate that our approach can be scaled up to systematically design and deliver highly efficient sgRNAs for the tissue-specific investigation of gene functions in Ciona.
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Registered trials
Read under generation 80e0d062 · epoch 390. Bibliography from PubMed, PubMed Central and OpenAlex; grants from NIH RePORTER; trial links from ClinicalTrials.gov; estimates, votes and beliefs from the OpenQuestion graph.