The DOI matches its Crossref record; 29 of the 29 references it cites were found in a registry or an index.
Highly Parallel Profiling of Cas9 Variant Specificity
Open accessAbstract
Determining the off-target cleavage profile of programmable nucleases is an important consideration for any genome editing experiment, and a number of Cas9 variants have been reported that improve specificity. We describe here tagmentation-based tag integration site sequencing (TTISS), an efficient, scalable method for analyzing double-strand breaks (DSBs) that we apply in parallel to eight Cas9 variants across 59 targets. Additionally, we generated thousands of other Cas9 variants and screened for variants with enhanced specificity and activity, identifying LZ3 Cas9, a high specificity variant with a unique +1 insertion profile. This comprehensive comparison reveals a general trade-off between Cas9 activity and specificity and provides information about the frequency of generation of +1 insertions, which has implications for correcting frameshift mutations.
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“Although some Cas9 variants have comparable editing activity and specificity with SpCas9 at a few target sites, high-throughput analysis is still lacking.”
From Machine learning-based prediction models to guide the selection of Cas9 variants for efficient gene editing (2024) · cites this paper as [23]
Experimental — accuracy not yet measuredThe source backs part of the claim, not all of it — or backs it for a different population, condition or comparison.
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“It worth mentioning that in several previous reports, LZ3 Cas9, Sniper Cas9, and HiFi Cas9 had comparable editing activity to the SpCas9-WT in human cells or other animal embryos.”
From Machine learning-based prediction models to guide the selection of Cas9 variants for efficient gene editing (2024) · cites this paper as [23]
Experimental — accuracy not yet measuredThe source backs part of the claim, not all of it — or backs it for a different population, condition or comparison.
In the citing paper
“Although LZ3 Cas9 did not display a stable 1 bp site-directed insertion pattern, as shown in previous studies according to the DSB repair results”
From Machine learning-based prediction models to guide the selection of Cas9 variants for efficient gene editing (2024) · cites this paper as [23]
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Citations
Cites
Genome engineering using the CRISPR-Cas9 system
Ran, F. A., Hsu, P. D., Wright, J., Agarwala, V., Scott, D. A., & Zhang, F. (2013) · Nature Protocols · doi:10.1038/nprot.2013.143 · Cited by 11,133
Self-citationGenome-scale CRISPR-Cas9 knockout screening in human cells
Shalem, O., Sanjana, N. E., Hartenian, E., Shi, X., Scott, D. A., Mikkelsen, T. S., Heckl, D., Ebert, B. L., Root, D. E., Doench, J. G., et al. (2014) · Science · doi:10.1126/science.1247005 · Cited by 5,234
Self-citationSearch-and-replace genome editing without double-strand breaks or donor DNA
Anzalone, A. V., Randolph, P. B., Davis, J. R., Sousa, A. A., Koblan, L. W., Levy, J. M., Chen, P. J., Wilson, C., Newby, G. A., Raguram, A., et al. (2019) · Nature · doi:10.1038/s41586-019-1711-4 · Cited by 4,767
High-fidelity CRISPR-Cas9 nucleases with no detectable genome-wide off-target effects
Kleinstiver, B. P., Pattanayak, V., Prew, M. S., Tsai, S. Q., Nguyen, N. T., Zheng, Z., & Joung, J. K. (2016) · Nature · doi:10.1038/nature16526 · Cited by 2,673
Rationally engineered Cas9 nucleases with improved specificity
Slaymaker, I. M., Gao, L., Zetsche, B., Scott, D. A., Yan, W. X., & Zhang, F. (2016) · Science · doi:10.1126/science.aad5227 · Cited by 2,414
Self-citationGUIDE-seq enables genome-wide profiling of off-target cleavage by CRISPR-Cas nucleases
Tsai, S. Q., Zheng, Z., Nguyen, N. T., Liebers, M., Topkar, V. V., Thapar, V., Wyvekens, N., Khayter, C., Iafrate, A. J., Le, L. P., et al. (2015) · Nature Biotechnology · doi:10.1038/nbt.3117 · Cited by 2,268
Evolved Cas9 variants with broad PAM compatibility and high DNA specificity
Hu, J. H., Miller, S. M., Geurts, M. H., Tang, W., Chen, L., Sun, N., Zeina, C. M., Gao, X., Rees, H. A., Lin, Z., et al. (2018) · Nature · doi:10.1038/nature26155 · Cited by 1,552
Enhanced proofreading governs CRISPR-Cas9 targeting accuracy
Chen, J. S., Dagdas, Y. S., Kleinstiver, B. P., Welch, M. M., Sousa, A. A., Harrington, L. B., Sternberg, S. H., Joung, J. K., Yildiz, A., & Doudna, J. A. (2017) · Nature · doi:10.1038/nature24268 · Cited by 1,168
Digenome-seq: Genome-wide profiling of CRISPR-cas9 off-target effects in human cells
Kim, D., Bae, S., Park, J., Kim, E., Kim, S., Yu, H. R., Hwang, J., Kim, J.-I., & Kim, J.-S. (2015) · Nature Methods · doi:10.1038/nmeth.3284 · Cited by 992
Tn5 transposase and tagmentation procedures for massively scaled sequencing projectsPreprint
Picelli, S., Björklund, Å. K., Reinius, B., Sagasser, S., Winberg, G., & Sandberg, R. (2014) · Genome Research · doi:10.1101/gr.177881.114 · Cited by 829
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Genome editing with CRISPR–Cas nucleases, base editors, transposases and prime editors
Anzalone, A. V., Koblan, L. W., & Liu, D. R. (2020) · Nature Biotechnology · doi:10.1038/s41587-020-0561-9 · Cited by 2,237
Past, present, and future of CRISPR genome editing technologies
Pacesa, M., Pelea, O., & Jinek, M. (2024) · Cell · doi:10.1016/j.cell.2024.01.042 · Cited by 609
The CRISPR-Cas toolbox and gene editing technologies
Liu, G., Lin, Q., Jin, S., & Gao, C. (2021) · Molecular Cell · doi:10.1016/j.molcel.2021.12.002 · Cited by 422
Gaussian accelerated molecular dynamics: Principles and applications
Wang, J., Arantes, P. R., Bhattarai, A., Hsu, R. V., Pawnikar, S. P., Huang, Y. M., Palermo, G., & Miao, Y. (2021) · Wiley Interdisciplinary Reviews Computational Molecular Science · doi:10.1002/wcms.1521 · Cited by 302
Kumar, R., Das, S. P., Choudhury, B. U., Kumar, A., Prakash, N. R., Verma, R., Chakraborti, M., Devi, A. G., Bhattacharjee, B., Das, R., et al. (2024) · Biological Research · doi:10.1186/s40659-024-00562-6 · Cited by 286
Metadata
- Authors
- Schmid-Burgk, J. L., Gao, L., Li, D., Gardner, Z., Strecker, J., Lash, B., & Zhang, F.
- Affiliations
- Broad Institute (US)
- Corresponding author
- Zhang, Feng
- Venue
- Molecular Cell
- Publisher
- Elsevier BV
- Published
- 2020-05
- Volume
- 78
- Issue
- 4
- Pages
- 794-800.e8
- ISSN
- 1097-2765 (ISSN-L)
- PMC
- PMC7370240
- OpenAlex
- W3011375666
- Cited by
- 197
- References
- 29
- Citations by year
- 2026: 11 · 2025: 25 · 2024: 42 · 2023: 34 · 2022: 38 · 2021: 34 · 2020: 13
- FWCI
- 9.65
- Type
- journal article
- Language
- en
- Open access
- bronze
- doi.org
- Licence
- Locations
- Molecular Cell (publishedVersion)
- PubMed (publishedVersion)
- PubMed Central (acceptedVersion)
- DSpace@MIT (Massachusetts Institute of Technology) (submittedVersion, cc-by-nc-nd)
- Funders
- National Institutes of Health
- G. Harold and Leila Y. Mathers Foundation
- Howard Hughes Medical Institute
- Edward Mallinckrodt, Jr. Foundation
- Topics
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › CRISPR and Genetic Engineering
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › RNA regulation and disease
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › DNA Repair Mechanisms
- Keywords
- Cas9 variants, off-target cleavage, DNA double-strand breaks, genome editing specificity, programmable nucleases
- MeSH terms
- Gene Editing, CRISPR-Associated Protein 9, RNA, Guide, CRISPR-Cas Systems, Humans, Genetic Variation, RNA, Guide, Kinetoplastida, K562 Cells, DNA Breaks, Double-Stranded, HEK293 Cells, CRISPR-Cas Systems
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