ArticleNucleic acids research2015
Development of an intein-mediated split-Cas9 system for gene therapy.
Article in Nucleic acids research, 2015. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 215 papers.
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Who cites it
215 citing papers in PubMed, 373 citations in OpenAlex.
- Lipid nanoparticles optimized for large RNA cargo and tissue targeting enhance in vivo genome editing.Nature biotechnology · 2026Article
- DNA Methylation as a Programmable Information Layer: From Molecular Marks to Disease State Engineering.International journal of molecular sciences · 2026Review
- Application of CRISPR-Cas9-Based Gene Editing Technology in Inherited Liver Diseases.International journal of molecular sciences · 2026Review
- Adapting prime editing with split prime editors in Escherichia coli and its application to Staphylococcus aureus genome editing.Applied microbiology and biotechnology · 2026Article
- Inducible CRISPR/Cas systems in precision oncology: Current applications and future perspectives.Clinical and translational medicine · 2026Review
- Considerations for early life genetic therapies in cystic fibrosis.American journal of physiology. Lung cellular and molecular physiology · 2026Review
- Prime editing updates: technological evolution, methodological expansion, and delivery strategies for in vivo applications.BMB reports · 2026Review
- CHARIOT-AAV: Conjugation of diverse vectors to adeno-associated viruses for delivery of large genes.bioRxiv : the preprint server for biology · 2026Article
- mRNA-engineered CRISPR-Cas epigenetic editors enable durable and efficient gene silencingInnovation (Cambridge (Mass.)) · 2026Article
- Advances and challenges of precision epigenetic therapy in treating genomic imprinting diseases.Translational pediatrics · 2026Review
- High-efficiency TadA cytosine base editors for precise modelling of human disease variants.Nature biomedical engineering · 2026Article
- Article
- Engineering TnpB as a compact RNA-guided genome editor from molecular constraints to design principles.Frontiers in genome editing · 2026Review
- Therapeutic in vivo genome editing: innovations and challenges in rAAV vector-based CRISPR delivery.Gene therapy · 2026Review
- Dystrophin Restorative and Compensatory Gene Addition Therapies for Duchenne Muscular Dystrophy: Could CRISPRa Provide a Realistic Alternative?Muscles (Basel, Switzerland) · 2025Review
- Engineering adeno-associated viral vectors for CRISPR/Cas based in vivo therapeutic genome editing.Biomaterials · 2025Review
- Optimized in vivo base editing restores auditory function in a DFNA15 mouse model.Nature communications · 2025Article
- Photoswitchable intein for light control of covalent protein binding and cleavage.Nature communications · 2025Article
- Trends and challenges of AAV-delivered gene editing therapeutics for CNS disorders: Implications for neurodegenerative disease.Molecular therapy. Nucleic acids · 2025Review
- Review
155 more citing papers are in PubMed but not listed here.
Corrections and comments
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Authors and funding
7 authors at 4 institutions in 1 country.
Funding
No grant is acknowledged in the PubMed record.
Abstract
Using CRISPR/Cas9, it is possible to target virtually any gene in any organism. A major limitation to its application in gene therapy is the size of Cas9 (>4 kb), impeding its efficient delivery via recombinant adeno-associated virus (rAAV). Therefore, we developed a split-Cas9 system, bypassing the packaging limit using split-inteins. Each Cas9 half was fused to the corresponding split-intein moiety and, only upon co-expression, the intein-mediated trans-splicing occurs and the full Cas9 protein is reconstituted. We demonstrated that the nuclease activity of our split-intein system is comparable to wild-type Cas9, shown by a genome-integrated surrogate reporter and by targeting three different endogenous genes. An analogously designed split-Cas9D10A nickase version showed similar activity as Cas9D10A. Moreover, we showed that the double nick strategy increased the homologous directed recombination (HDR). In addition, we explored the possibility of delivering the repair template accommodated on the same dual-plasmid system, by transient transfection, showing an efficient HDR. Most importantly, we revealed for the first time that intein-mediated split-Cas9 can be packaged, delivered and its nuclease activity reconstituted efficiently, in cells via rAAV.
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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.