ArticleNature biotechnology2023
High-yield genome engineering in primary cells using a hybrid ssDNA repair template and small-molecule cocktails.
Article in Nature biotechnology, 2023. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 136 papers.
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Who cites it
136 citing papers in PubMed, 207 citations in OpenAlex.
- Chromatin Modifications Orchestrate DNA Repair Pathway Choice: Epigenetics Strategies for Enhancing CRISPR-Cas9-Mediated Knock-In Efficiency.Cell proliferation · 2026Review
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- Double-stranded DNA donors and CRISPR-Cas9 for universal correction of mutations causing cystic fibrosis in human airway cells.Molecular therapy. Nucleic acids · 2026Article
- Integrated epigenetic and genetic programming of primary human T cells.Nature biotechnology · 2026Article
- Article
- From Delivery to Design: Site-Specific Genome Engineering for Next-Generation CAR T-Cell Therapy.Biomedicines · 2026Review
- Virus-like particles enable targeted gene engineering and pooled CRISPR screening in primary human myeloid cells.Nature biotechnology · 2026Article
- Review
- The T Cell Receptor: Molecular Sensor, Therapeutic Mediator and Probabilistic Driver of Adaptive Immunity.Immunological reviews · 2026Review
- Prime editing-mediated microhomology enables efficient replacement of large DNA.Nucleic acids research · 2026Article
- Article
- Modeling and correction of SCID-X1 using CRISPR-Cas9 homology-directed repair in human HSPCs.Molecular therapy. Nucleic acids · 2026Article
- Programmable Nucleic Acid Sensing in Human Cells Using Circularizable ssDNA.Nature communications · 2026Article
- Dual CD8 and TCR editing in regulatory T cells mediates HLA-A2-restricted tissue-specific homing.Molecular therapy : the journal of the American Society of Gene Therapy · 2026Article
- Precise, predictable genome integrations by deep-learning-assisted design of microhomology-based templates.Nature biotechnology · 2026Article
- Article
- Selection of human hematopoietic stem cells bearing the intended functional edit by transient AND-gate reporters.Nature biotechnology · 2026Article
- A bulk cell heterozygous knock-in strategy for targeted protein degradation.bioRxiv : the preprint server for biology · 2026Article
- Simultaneous orthogonal cell engineering by a single CRISPR-Cas9 polyfunctional editor.Nature communications · 2026Article
- Parallel Activation and Interference CRISPR (PAIR) with Sequencing Uncovers DNA Repair Networks Guiding Precision Cell Engineering.bioRxiv : the preprint server for biology · 2026Article
76 more citing papers are in PubMed but not listed here.
Corrections and comments
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Authors and funding
23 authors at 3 institutions in 2 countries.
Funding
Abstract
Enhancing CRISPR-mediated site-specific transgene insertion efficiency by homology-directed repair (HDR) using high concentrations of double-stranded DNA (dsDNA) with Cas9 target sequences (CTSs) can be toxic to primary cells. Here, we develop single-stranded DNA (ssDNA) HDR templates (HDRTs) incorporating CTSs with reduced toxicity that boost knock-in efficiency and yield by an average of around two- to threefold relative to dsDNA CTSs. Using small-molecule combinations that enhance HDR, we could further increase knock-in efficiencies by an additional roughly two- to threefold on average. Our method works across a variety of target loci, knock-in constructs and primary human cell types, reaching HDR efficiencies of >80-90%. We demonstrate application of this approach for both pathogenic gene variant modeling and gene-replacement strategies for IL2RA and CTLA4 mutations associated with Mendelian disorders. Finally, we develop a good manufacturing practice (GMP)-compatible process for nonviral chimeric antigen receptor-T cell manufacturing, with knock-in efficiencies (46-62%) and yields (>1.5 × 10
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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.