The DOI matches its Crossref record; 29 of the 29 references it cites were found in a registry or an index.
DNA targeting specificity of RNA-guided Cas9 nucleases
Open accessAbstract
The Streptococcus pyogenes Cas9 (SpCas9) nuclease can be efficiently targeted to genomic loci by means of single-guide RNAs (sgRNAs) to enable genome editing. Here, we characterize SpCas9 targeting specificity in human cells to inform the selection of target sites and avoid off-target effects. Our study evaluates >700 guide RNA variants and SpCas9-induced indel mutation levels at >100 predicted genomic off-target loci in 293T and 293FT cells. We find that SpCas9 tolerates mismatches between guide RNA and target DNA at different positions in a sequence-dependent manner, sensitive to the number, position and distribution of mismatches. We also show that SpCas9-mediated cleavage is unaffected by DNA methylation and that the dosage of SpCas9 and sgRNA can be titrated to minimize off-target modification. To facilitate mammalian genome engineering applications, we provide a web-based software tool to guide the selection and validation of target sequences as well as off-target analyses.
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“For example, off-target cleavages of the Cas9 nucleases that cause oncogenic mutations in the genome have been reported extensively [123,124].”
From Utility of iPSC-Derived Cells for Disease Modeling, Drug Development, and Cell Therapy (2022) · cites this paper as [123]
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
“The off-target sites are often sgRNA-dependent, since Cas9 is known to tolerate up to 3 mismatches between sgRNA and genomic DNA (Fu et al., 2013; Hsu et al., 2013; Wang et al., 2016a).”
From Off-target effects in CRISPR/Cas9 gene editing (2023) · cites this paper as Hsu et al., 2013
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 CRISPR/Cas systems exhibit tremendous potential in translational medicine, off-target effects remain a major challenge (Fu et al., 2013; Hsu et al., 2013; Pacesa et al., 2022).”
From Off-target effects in CRISPR/Cas9 gene editing (2023) · cites this paper as Hsu et al., 2013
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
“and it maintains cleaving activity even with 3-5 mismatches at the PAM-distal end of the gRNA molecule [59,81,102].”
From CRISPR-Cas: Converting A Bacterial Defence Mechanism into A State-of-the-Art Genetic Manipulation Tool (2019) · cites this paper as [102]
Experimental — accuracy not yet measuredThe cited source says what the claim attributes to it.
In the citing paper
“The algorithm of MIT weights the position effect of the mismatches in the sgRNA to generate a score to evaluate off-target effects (Hsu et al., 2013; Haeussler et al., 2016).”
From Off-target effects in CRISPR/Cas9 gene editing (2023) · cites this paper as Hsu et al., 2013
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“The PAM sequence adjacent to the target sequence greatly affects the editing efficiency and editing range of Cas9.”
From Machine learning-based prediction models to guide the selection of Cas9 variants for efficient gene editing (2024) · cites this paper as [2]
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In the citing paper
“Cas9 is guided by a 20 nt fraction of the gRNA”
From CRISPR-Cas: Converting A Bacterial Defence Mechanism into A State-of-the-Art Genetic Manipulation Tool (2019) · cites this paper as [102]
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In the citing paper
“Recombinant expression and purification of EvoCas9–1 paired with chemically synthesized Evo-generated single guide RNA (sgRNA) exhibited comparable in vitro cleavage activity to SpCas9 paired with the canonical SpCas9 sgRNA”
From Sequence modeling and design from molecular to genome scale with Evo (2024) · cites this paper as [63]
Experimental — accuracy not yet measuredThe check wasn’t confident either way, so it gives no verdict — never disputed.
In the citing paper
“When two sgRNAs are designed to simultaneously cut the two opposite strands of DNA within a small distance, DSBs can be created with markedly lower off-target effects (Hsu et al., 2013; Frock et al., 2015).”
From Off-target effects in CRISPR/Cas9 gene editing (2023) · cites this paper as Hsu et al., 2013
Experimental — accuracy not yet measuredIn the citing paper
“By bioinformatic screening and efficiency/off-target prediction (14, 15), we identified four gRNA targets (protospacers; LTRs A-D) that avoid conserved transcription factor binding sites, minimizing the likelihood of altering host gene expression”
From RNA-directed gene editing specifically eradicates latent and prevents new HIV-1 infection (2014) · cites this paper as [14]
In the citing paper
“as mentioned in several recent publications, Cas9 tolerates mismatches between the guide RNA and target DNA in a sequence-dependent manner and potentially tolerates up to five target mismatches in human cells [12], [24], [25], [26], [27], thereby promoting undesired off-target mutations.”
From High-efficiency targeted editing of large viral genomes by RNA-guided nucleases (2014) · cites this paper as [25]
In the citing paper
“Because the sequence specificity of Streptococcus pyogenes Cas9 tolerates 4 mismatches between the gRNA and complementary sequence”
From High-efficiency targeted editing of large viral genomes by RNA-guided nucleases (2014) · cites this paper as [25]
In the citing paper
“non-specific cleavage may occur in the cellular genome [12], [24], [25], [26], [27].”
From High-efficiency targeted editing of large viral genomes by RNA-guided nucleases (2014) · cites this paper as [25]
In the citing paper
“However, it has been recently shown that CRISPR/Cas9 system can be highly active even when mismatches occur between gRNA and their DNA targets”
From Newer Gene Editing Technologies toward HIV Gene Therapy (2013) · cites this paper as [91]
In the citing paper
“multiple mismatches between the guide RNA and its complementary target DNA sequence can be tolerated depending on the quantity, position, and base identity of mismatches [87,90,91,93], all of which contribute to potential off-target activity.”
From Newer Gene Editing Technologies toward HIV Gene Therapy (2013) · cites this paper as [91]
In the citing paper
“and off-target mutations occur at endogenous loci in human cells with high frequency [90,91,92].”
From Newer Gene Editing Technologies toward HIV Gene Therapy (2013) · cites this paper as [91]
In the citing paper
“Collectively, these studies indicate that although each base within the 20 nt guide sequence contributes to overall specificity”
From Newer Gene Editing Technologies toward HIV Gene Therapy (2013) · cites this paper as [91]
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Citations
Cites
A Programmable Dual-RNA–Guided DNA Endonuclease in Adaptive Bacterial Immunity
Jinek, M., Chylinski, K., Fonfara, I., Hauer, M., Doudna, J. A., & Charpentier, E. (2012) · Science · doi:10.1126/science.1225829 · Cited by 17,116
Multiplex Genome Engineering Using CRISPR/Cas Systems
Cong, L., Ran, F. A., Cox, D., Lin, S., Barretto, R., Habib, N., Hsu, P. D., Wu, X., Jiang, W., Marraffini, L. A., et al. (2013) · Science · doi:10.1126/science.1231143 · Cited by 15,339
Self-citationRNA-Guided Human Genome Engineering via Cas9
Mali, P., Yang, L., Esvelt, K. M., Aach, J., Guell, M., DiCarlo, J. E., Norville, J. E., & Church, G. M. (2013) · Science · doi:10.1126/science.1232033 · Cited by 9,210
Repurposing CRISPR as an RNA-Guided Platform for Sequence-Specific Control of Gene Expression
Qi, L. S., Larson, M. H., Gilbert, L. A., Doudna, J. A., Weissman, J. S., Arkin, A. P., & Lim, W. A. (2013) · Cell · doi:10.1016/j.cell.2013.02.022 · Cited by 5,016
Wang, H., Yang, H., Shivalila, C. S., Dawlaty, M. M., Cheng, A. W., Zhang, F., & Jaenisch, R. (2013) · Cell · doi:10.1016/j.cell.2013.04.025 · Cited by 3,495
Self-citationEfficient genome editing in zebrafish using a CRISPR-Cas system
Hwang, W. Y., Fu, Y., Reyon, D., Maeder, M. L., Tsai, S. Q., Sander, J. D., Peterson, R. T., Yeh, J.-R. J., & Joung, J. K. (2013) · Nature Biotechnology · doi:10.1038/nbt.2501 · Cited by 2,937
CRISPR RNA Maturation by Trans-Encoded Small RNA and Host Factor RNase III
Deltcheva, E., Chylinski, K., Sharma, C. M., Gonzales, K., Chao, Y., Pirzada, Z. A., Eckert, M. R., Vogel, J., & Charpentier, E. (2011) · Nature · doi:10.1038/nature09886 · Cited by 2,563
RNA-guided editing of bacterial genomes using CRISPR-Cas systems
Jiang, W., Bikard, D., Cox, D., Zhang, F., & Marraffini, L. A. (2013) · Nature Biotechnology · doi:10.1038/nbt.2508 · Cited by 2,429
Self-citationA TALE nuclease architecture for efficient genome editing
Miller, J. C., Tan, S., Qiao, G., Barlow, K. A., Wang, J., Xia, D. F., Meng, X., Paschon, D. E., Leung, E., Hinkley, S. J., et al. (2011) · Nature Biotechnology · doi:10.1038/nbt.1755 · Cited by 2,098
RNA-programmed genome editing in human cells
Jinek, M., East, A., Cheng, A., Lin, S., Ma, E., & Doudna, J. (2013) · eLife · doi:10.7554/elife.00471 · Cited by 2,098
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Ausländer, S., Ausländer, D., & Fussenegger, M. (2016) · Angewandte Chemie International Edition · doi:10.1002/anie.201609229 · Cited by 210
Modern Trends in Plant Genome Editing: An Inclusive Review of the CRISPR/Cas9 Toolbox
Razzaq, A., Saleem, F., Kanwal, M., Mustafa, G., Yousaf, S., Arshad, H. M. I., Hameed, M. K., Khan, M. S., & Joyia, F. A. (2019) · International Journal of Molecular Sciences · doi:10.3390/ijms20164045 · Cited by 216
Evaluation of TCR Gene Editing Achieved by TALENs, CRISPR/Cas9, and megaTAL Nucleases
Osborn, M. J., Webber, B. R., Knipping, F., Lonetree, C.-L., Tennis, N., DeFeo, A. P., McElroy, A. N., Starker, C. G., Lee, C., Merkel, S., et al. (2016) · Molecular Therapy · doi:10.1038/mt.2015.197 · Cited by 201
Li, J., Manghwar, H., Sun, L., Wang, P., Wang, G., Sheng, H., Zhang, J., Liu, H., Qin, L., Rui, H., et al. (2018) · Plant Biotechnology Journal · doi:10.1111/pbi.13020 · Cited by 211
CRISPR/Cas-mediated genome editing in the rat via direct injection of one-cell embryos
Shao, Y., Guan, Y., Wang, L., Qiu, Z., Liu, M., Chen, Y., Wu, L., Li, Y., Ma, X., Liu, M., et al. (2014) · Nature Protocols · doi:10.1038/nprot.2014.171 · Cited by 207
Self-citationThe CRISPR–Cas system for plant genome editing: advances and opportunities
Kumar, V. S., & Jain, M. (2014) · Journal of Experimental Botany · doi:10.1093/jxb/eru429 · Cited by 207
Boyle, E. A., Andreasson, J. O. L., Chircus, L. M., Sternberg, S. H., Wu, M. J., Guegler, C. K., Doudna, J. A., & Greenleaf, W. J. (2017) · Proceedings of the National Academy of Sciences · doi:10.1073/pnas.1700557114 · Cited by 205
Easy quantification of template-directed CRISPR/Cas9 editing
Brinkman, E. K., Kousholt, A. N., Harmsen, T., Leemans, C., Chen, T., Jonkers, J., & van Steensel, B. (2018) · Nucleic Acids Research · doi:10.1093/nar/gky164 · Cited by 197
Profiling of engineering hotspots identifies an allosteric CRISPR-Cas9 switch
Oakes, B. L., Nadler, D. C., Flamholz, A. I., Fellmann, C., Staahl, B. T., Doudna, J. A., & Savage, D. F. (2016) · Nature Biotechnology · doi:10.1038/nbt.3528 · Cited by 199
Computational approaches for effective CRISPR guide RNA design and evaluation
Liu, G., Zhang, Y., & Zhang, T. (2019) · Computational and Structural Biotechnology Journal · doi:10.1016/j.csbj.2019.11.006 · Cited by 206
“The CRISPR/Cas system was first adapted for gene editing in 2012, and several tools were developed immediately thereafter, such as Zinc Finger Targeter (ZiFiT) [89], Cas9 guide RNA Design [68] and CRISPR (http://crispr.mit.edu/) [41].”
BackgroundMethodInfluentialSequence features associated with the cleavage efficiency of CRISPR/Cas9 system
Liu, X., Homma, A., Sayadi, J., Yang, S., Ohashi, J., & Takumi, T. (2016) · Scientific Reports · doi:10.1038/srep19675 · Cited by 166
Efficiency and Inheritance of Targeted Mutagenesis in Maize Using CRISPR-Cas9
Zhu, J., Song, N., Sun, S., Yang, W., Zhao, H., Song, W., & Lai, J. (2015) · Journal of genetics and genomics/Journal of Genetics and Genomics · doi:10.1016/j.jgg.2015.10.006 · Cited by 203
Prokaryotic Argonaute proteins: novel genome-editing tools?
Hegge, J. W., Swarts, D. C., & John van der, O. O. S. T. (2017) · Nature Reviews Microbiology · doi:10.1038/nrmicro.2017.73 · Cited by 199
Abadi, S., Yan, W. X., Amar, D., & Mayrose, I. (2017) · PLoS Computational Biology · doi:10.1371/journal.pcbi.1005807 · Cited by 208
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Song, C.-Q., Jiang, T., Richter, M., Rhym, L. H., Koblan, L. W., Zafra, M. P., Schatoff, E. M., Doman, J. L., Cao, Y., Dow, L. E., et al. (2019) · Nature Biomedical Engineering · doi:10.1038/s41551-019-0357-8 · Cited by 200
Comparison of non-canonical PAMs for CRISPR/Cas9-mediated DNA cleavage in human cells
Zhang, Y., Ge, X., Yang, F., Zhang, L., Zheng, J., Tan, X., Jin, Z., Qu, J., & Gu, F. (2014) · Scientific Reports · doi:10.1038/srep05405 · Cited by 205
Strategies for nonviral nanoparticle‐based delivery of CRISPR/Cas9 therapeutics
Chen, F., Alphonse, M., & Liu, Q. (2020) · WIREs Nanomedicine and Nanobiotechnology · doi:10.1002/wnan.1609 · Cited by 197
Lee, J. S., Kallehauge, T. B., Pedersen, L. E., & Kildegaard, H. F. (2015) · Scientific Reports · doi:10.1038/srep08572 · Cited by 203
A time-resolved, multi-symbol molecular recorder via sequential genome editing
Choi, J., Chen, W., Minkina, A., Chardon, F. M., Suiter, C. C., Regalado, S. G., Domcke, S., Hamazaki, N., Lee, C., Martin, B., et al. (2022) · Nature · doi:10.1038/s41586-022-04922-8 · Cited by 202
Gao, X., Tsang, J. C. H., Gaba, F., Wu, D., Lu, L., & Liu, P. (2014) · Nucleic Acids Research · doi:10.1093/nar/gku836 · Cited by 200
Zhu, L. J., Holmes, B. R., D, N. A. M., & Brodsky, M. H. (2014) · PLoS ONE · doi:10.1371/journal.pone.0108424 · Cited by 192
Smith, J., Suresh, S., Schlecht, U., Wu, M., Wagih, O., Peltz, G., Davis, R. W., Steinmetz, L. M., Parts, L., & St.Onge, R. P. (2016) · Genome biology · doi:10.1186/s13059-016-0900-9 · Cited by 199
Inherent DNA‐binding specificities of the HIF‐1α and HIF‐2α transcription factors in chromatin
Smythies, J. A., Sun, M., Masson, N., Salama, R., Simpson, P. D., Murray, E., Neumann, V., Cockman, M. E., Choudhry, H., Ratcliffe, P. J., et al. (2019) · The EMBO Reports · doi:10.15252/embr.201846401 · Cited by 195
Smith, I. C. P., Greenside, P., Natoli, T. E., Lahr, D. L., Wadden, D., Tirosh, I., Narayan, R., Root, D. E., Golub, T. R., Subramanian, A., et al. (2017) · PLoS Biology · doi:10.1371/journal.pbio.2003213 · Cited by 197
Hypoxia inducible factor-1α drives cancer resistance to cuproptosis
Yang, Z., Su, W., Wei, X., Pan, Y., Xing, M., Niu, L., Feng, B., Kong, W., Ren, X., Huang, F., et al. (2025) · Cancer Cell · doi:10.1016/j.ccell.2025.02.015 · Cited by 208
Chen, X., Gao, C., Guo, L., Hu, G., Luo, Q., Liu, J., Nielsen, J., Chen, J., & Liu, L. (2017) · Chemical Reviews · doi:10.1021/acs.chemrev.6b00804 · Cited by 191
Targeted mutagenesis in the medicinal plant Salvia miltiorrhiza
Li, B., Cui, G., Shen, G., Zhan, Z.-L., Huang, L., Chen, J., & Qi, X. (2017) · Scientific Reports · doi:10.1038/srep43320 · Cited by 191
A CRISPR toolbox to study virus–host interactions
Puschnik, A. S., Majzoub, K., Ooi, Y. S., & Carette, J. E. (2017) · Nature Reviews Microbiology · doi:10.1038/nrmicro.2017.29 · Cited by 185
György, B., Lööv, C., Zaborowski, M. P., Takeda, S., Kleinstiver, B. P., Commins, C., Kastanenka, K. V., Mu, D., Volak, A., Giedraitis, V., et al. (2018) · Molecular Therapy — Nucleic Acids · doi:10.1016/j.omtn.2018.03.007 · Cited by 205
Ronda, C., Pedersen, L. E., Hansen, H. G., Kallehauge, T. B., Betenbaugh, M. J., Nielsen, A. T., & Kildegaard, H. F. (2014) · Biotechnology and Bioengineering · doi:10.1002/bit.25233 · Cited by 195
Šťovíček, V., Borodina, I., & Förster, J. (2015) · Metabolic Engineering Communications · doi:10.1016/j.meteno.2015.03.001 · Cited by 190
Delivery and Specificity of CRISPR/Cas9 Genome Editing Technologies for Human Gene Therapy
Gori, J. L., Hsu, P. D., Maeder, M. L., Shen, S., Welstead, G. G., & Bumcrot, D. A. (2015) · Human Gene Therapy · doi:10.1089/hum.2015.074 · Cited by 200
Self-citationCRISPy-web: An online resource to design sgRNAs for CRISPR applications
Blin, K., Pedersen, L. E., Weber, T., & Lee, S. Y. (2016) · Synthetic and Systems Biotechnology · doi:10.1016/j.synbio.2016.01.003 · Cited by 185
TALENs facilitate targeted genome editing in human cells with high specificity and low cytotoxicity
Mussolino, C., Alzubi, J., Fine, E. J., Morbitzer, R., Cradick, T. J., Lahaye, T., Bao, G., & Cathomen, T. (2014) · Nucleic Acids Research · doi:10.1093/nar/gku305 · Cited by 194
Self-citationMagnetic iron oxide nanoparticles for disease detection and therapy
Tong, S., Zhu, H., & Bao, G. (2019) · Materials Today · doi:10.1016/j.mattod.2019.06.003 · Cited by 193
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Vats, S., Kumawat, S., Kumar, V., Patil, G., Joshi, T., Sonah, H., Sharma, T. R., & Deshmukh, R. K. (2019) · Cells · doi:10.3390/cells8111386 · Cited by 195
“Reports suggest that Cas9/sgRNA cannot recognize and edit a DNA site with mismatches within 10–12 bp of the PAM site, and the Cas9 protein has a higher affinity for NGG-PAM than for NAG-PAM [155].”
BackgroundZhang, Y., Qian, L., Wei, W., Wang, Y., Wang, B., Lin, P., Liu, W., Xu, L., Li, X., Liu, D., et al. (2016) · ACS Synthetic Biology · doi:10.1021/acssynbio.6b00215 · Cited by 189
Zheng, M., Zhang, L. N., Tang, M., Liu, J., Liu, H., Yang, H., Fan, S., Terzaghi, W., Wang, H., & Hua, W. (2019) · Plant Biotechnology Journal · doi:10.1111/pbi.13228 · Cited by 196
The evolving landscape around genome editing in agriculture
Schmidt, S. M., Belisle, M., & Frommer, W. B. (2020) · EMBO Reports · doi:10.15252/embr.202050680 · Cited by 189
CRISPR/Cas9-mediated targeted chromosome elimination
Zuo, E., Huo, X., Yao, X., Hu, X., Sun, Y., Yin, J., He, B., Wang, X., Shi, L., Ping, J., et al. (2017) · Genome biology · doi:10.1186/s13059-017-1354-4 · Cited by 194
Self-citationMurovec, J., Guček, K., Bohanec, B., Avbelj, M., & Jerala, R. (2018) · Frontiers in Plant Science · doi:10.3389/fpls.2018.01594 · Cited by 189
Efficient gene editing of human long-term hematopoietic stem cells validated by clonal tracking
Ferrari, S., Jacob, A., Beretta, S., Unali, G., Albano, L., Vavassori, V., Cittaro, D., Lazarević, D., Brombin, C., Cugnata, F., et al. (2020) · Nature Biotechnology · doi:10.1038/s41587-020-0551-y · Cited by 188
“Sequences of the gRNAs were designed using an online tool 54 and selected for predicted specificity score and on-target activity.”
Höijer, I., Emmanouilidou, A., Östlund, R., van Schendel, R., Bozorgpana, S., Tijsterman, M., Feuk, L., Gyllensten, U. B., den Hoed, M., & Ameur, A. (2022) · Nature Communications · doi:10.1038/s41467-022-28244-5 · Cited by 205
Chen, B., Hu, J., Almeida, R., Liu, H., Balakrishnan, S., Covill‐Cooke, C., Lim, W. A., & Huang, B. (2016) · Nucleic Acids Research · doi:10.1093/nar/gkv1533 · Cited by 190
Bothmer, A., Phadke, T., Barrera, L. A., Margulies, C. M., Lee, C. S., Buquicchio, F., Moss, S., Abdulkerim, H. S., Selleck, W., Jayaram, H., et al. (2017) · Nature Communications · doi:10.1038/ncomms13905 · Cited by 187
Zhang, Q., Xing, H.-L., Wang, Z.-P., Zhang, H., Yang, F., Wang, X., & Chen, Q. (2018) · Plant Molecular Biology · doi:10.1007/s11103-018-0709-x · Cited by 195
Engineering of high-precision base editors for site-specific single nucleotide replacement
Tan, J., Zhang, F., Karcher, D., & Bock, R. (2019) · Nature Communications · doi:10.1038/s41467-018-08034-8 · Cited by 198
Self-citationSynthetic CRISPR RNA-Cas9–guided genome editing in human cells
Rahdar, M., McMahon, M. A., Prakash, T. P., Swayze, E. E., Bennett, C. F., & Cleveland, D. W. (2015) · Proceedings of the National Academy of Sciences · doi:10.1073/pnas.1520883112 · Cited by 184
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Song, R., Zhai, Q., Sun, L., Huang, E., Zhang, Y., Zhu, Y., Guo, Q., Tian, Y., Zhao, B., & Lu, H. X. (2019) · Applied Microbiology and Biotechnology · doi:10.1007/s00253-019-10007-w · Cited by 194
Hultquist, J. F., Schumann, K., Woo, J. M., Manganaro, L., McGregor, M. J., Doudna, J., Simon, V., Krogan, N. J., & Marson, A. (2016) · Cell Reports · doi:10.1016/j.celrep.2016.09.080 · Cited by 170
Off-target predictions in CRISPR-Cas9 gene editing using deep learning
Lin, J., & Wong, K.-C. (2018) · Bioinformatics · doi:10.1093/bioinformatics/bty554 · Cited by 185
Precision genome editing in the CRISPR era
Salsman, J., & Dellaire, G. (2016) · Biochemistry and Cell Biology · doi:10.1139/bcb-2016-0137 · Cited by 193
CRISPR-SKIP: programmable gene splicing with single base editors
Gapinske, M., Luu, A., Winter, J., Woods, W. S., Kostan, K. A., Shiva, N., Song, J. S., & Pérez‐Piñera, P. (2018) · Genome biology · doi:10.1186/s13059-018-1482-5 · Cited by 191
Peterson, B. A., Haak, D. C., Nishimura, M. T., Teixeira, P. J. P. L., James, S. R., Dangl, J. L., & Nimchuk, Z. L. (2016) · PLOS ONE · doi:10.1371/journal.pone.0162169 · Cited by 172
Marker-free coselection for CRISPR-driven genome editing in human cells
Agudelo, D., Duringer, A., Bozoyan, L., Huard, C., Carter, S., Loehr, J., Synodinou, D., Drouin, M., Salsman, J., Dellaire, G., et al. (2017) · Nature Methods · doi:10.1038/nmeth.4265 · Cited by 183
RNA-dependent RNA targeting by CRISPR-Cas9
Strutt, S. C., Torrez, R. M., Kaya, E., Negrete, O. A., & Doudna, J. A. (2018) · eLife · doi:10.7554/elife.32724 · Cited by 186
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Blitz, I. L., Biesinger, J., Xie, X. S., & Cho, K. W. Y. (2013) · genesis · doi:10.1002/dvg.22719 · Cited by 185
TALE nickase-mediated SP110 knockin endows cattle with increased resistance to tuberculosis
Wu, H., Wang, Y., Zhang, Y., Yang, M., Lv, J., Liu, J., & Zhang, Y. (2015) · Proceedings of the National Academy of Sciences · doi:10.1073/pnas.1421587112 · Cited by 185
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Zhang, F. (2019) · Quarterly Reviews of Biophysics · doi:10.1017/s0033583519000052 · Cited by 188
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Okada, A., Arndell, T., Borisjuk, N. V., Sharma, N., Watson‐Haigh, N. S., Tucker, E. J., Baumann, U., Langridge, P., & Whitford, R. (2019) · Plant Biotechnology Journal · doi:10.1111/pbi.13106 · Cited by 181
SHANK2 mutations associated with autism spectrum disorder cause hyperconnectivity of human neurons
Zaslavsky, K., Zhang, W.-B., McCready, F. P., Rodrigues, D. C., Deneault, E., Loo, C., Zhao, M., Ross, P. J., El Hajjar, J., Romm, A., et al. (2019) · Nature Neuroscience · doi:10.1038/s41593-019-0365-8 · Cited by 174
CRISPR/Cas9 Immune System as a Tool for Genome Engineering
Hryhorowicz, M., Lipiński, D., Zeyland, J., & Słomski, R. (2016) · Archivum Immunologiae et Therapiae Experimentalis · doi:10.1007/s00005-016-0427-5 · Cited by 206
Park, C.-Y., Halevy, T., Lee, D. R., Sung, J. J., Lee, J. S., Yanuka, O., Benvenisty, N., & Kim, D.-W. (2015) · Cell Reports · doi:10.1016/j.celrep.2015.08.084 · Cited by 178
Yoshimi, K., Kaneko, T., Voigt, B., & Mashimo, T. (2014) · Nature Communications · doi:10.1038/ncomms5240 · Cited by 178
CRISPR-Cas: an efficient tool for genome engineering of virulent bacteriophages
Martel, B., & Moineau, S. (2014) · Nucleic Acids Research · doi:10.1093/nar/gku628 · Cited by 182
Multidimensional chemical control of CRISPR–Cas9
Maji, B., Moore, C. L., Zetsche, B., Volz, S. E., Zhang, F., Shoulders, M. D., & Choudhary, A. (2016) · Nature Chemical Biology · doi:10.1038/nchembio.2224 · Cited by 180
Self-citationCRISPR/Cas9-mediated genome editing induces exon skipping by alternative splicing or exon deletion
Mou, H., Smith, J. L., Peng, L., Yin, H., Moore, J. E., Zhang, X.-O., Song, C., Sheel, A., Wu, Q., Özata, D. M., et al. (2017) · Genome biology · doi:10.1186/s13059-017-1237-8 · Cited by 190
Self-citational Amin, N., Ahmad, N., Wu, N., Pu, X., Ma, T., Du, Y., Bo, X., Wang, N., Sharif, R., & Wang, P. (2019) · BMC Biotechnology · doi:10.1186/s12896-019-0501-2 · Cited by 178
Qin, L., Li, J., Wang, Q., Xu, Z., Sun, L., Alariqi, M., Manghwar, H., Wang, G., Li, B., Ding, X. L., et al. (2019) · Plant Biotechnology Journal · doi:10.1111/pbi.13168 · Cited by 179
Li, Y., Zhang, J., Chen, D., Yang, P., Jiang, F., Wang, X., & Kang, L. (2016) · Insect Biochemistry and Molecular Biology · doi:10.1016/j.ibmb.2016.10.003 · Cited by 178
Self-citationThe genomic landscape of TERT promoter wildtype-IDH wildtype glioblastoma
Diplas, B. H., He, X., Brosnan‐Cashman, J. A., Liu, H., Chen, L. H., Wang, Z., Moure, C. J., Killela, P. J., Loriaux, D. B., Lipp, E., et al. (2018) · Nature Communications · doi:10.1038/s41467-018-04448-6 · Cited by 175
The Impact of Chromatin Dynamics on Cas9-Mediated Genome Editing in Human Cells
Daer, R., Cutts, J., Brafman, D. A., & Haynes, K. A. (2016) · ACS Synthetic Biology · doi:10.1021/acssynbio.5b00299 · Cited by 184
Li, W., Wang, K., Chern, M., Liu, Y., Zhu, Z., Liu, J., Zhu, X., Yin, J., Li, R., Xiong, J., et al. (2020) · New Phytologist · doi:10.1111/nph.16505 · Cited by 179
Self-citationEfficient and Allele-Specific Genome Editing of Disease Loci in Human iPSCs
Smith, C. J., Abalde-Atristain, L., He, C., Brodsky, B. R., Braunstein, E. M., Chaudhari, P., Jang, Y.-Y., Cheng, L., & Ye, Z. (2014) · Molecular Therapy · doi:10.1038/mt.2014.226 · Cited by 180
Ordon, J., Gantner, J., Kemna, J., Lennart, S., Reschke, M., Streubel, J., Boch, J., & Stuttmann, J. (2016) · The Plant Journal · doi:10.1111/tpj.13319 · Cited by 176
Prykhozhij, S. V., Rajan, V., Gaston, D., & Berman, J. N. (2015) · PLoS ONE · doi:10.1371/journal.pone.0119372 · Cited by 173
Fackler, N., Heijstra, B. D., Rasor, B. J., Brown, H., Martin, J. P., Ni, Z., Shebek, K. M., Rosin, R. R., Simpson, S. D., Tyo, K. E. J., et al. (2021) · Annual Review of Chemical and Biomolecular Engineering · doi:10.1146/annurev-chembioeng-120120-021122 · Cited by 176
CRISPR-Cas9D10A nickase-based genotypic and phenotypic screening to enhance genome editing
Chiang, T.-W. W., le Sage, C., Larrieu, D., Demir, M., & Jackson, S. P. (2016) · Scientific Reports · doi:10.1038/srep24356 · Cited by 171
CRISPR/Cas9: A powerful tool for crop genome editing
Song, G., Jia, M., Chen, K., Kong, X., Khattak, B., Xie, C., Li, A., & Mao, L. (2016) · The Crop Journal · doi:10.1016/j.cj.2015.12.002 · Cited by 180
CRISPR-Cas9 off-targeting assessment with nucleic acid duplex energy parameters
Alkan, F., Wenzel, A., Anthon, C., Havgaard, J. H., & Gorodkin, J. (2018) · Genome biology · doi:10.1186/s13059-018-1534-x · Cited by 174
Willems, J., de Jong, A. P. H., Scheefhals, N., Mertens, E. J., Catsburg, L. A. E., Poorthuis, R. B., de Winter, F., Verhaagen, J., Meye, F. J., & MacGillavry, H. D. (2020) · PLoS Biology · doi:10.1371/journal.pbio.3000665 · Cited by 176
R-loop formation and conformational activation mechanisms of Cas9
Pačesa, M., Loeff, L., Querques, I., Muckenfuss, L. M., Sawicka, M., & Jínek, M. (2022) · Nature · doi:10.1038/s41586-022-05114-0 · Cited by 176
Evaluating and enhancing target specificity of gene-editing nucleases and deaminases
Kim, D., Luk, K., Wolfe, S. A., & Kim, J.-S. (2019) · Annual Review of Biochemistry · doi:10.1146/annurev-biochem-013118-111730 · Cited by 171
CRISPR/Cas9 and TALEN-mediated knock-in approaches in zebrafish
Auer, T. O., & Bene, F. D. (2014) · Methods · doi:10.1016/j.ymeth.2014.03.027 · Cited by 178
Genome Editing—Principles and Applications for Functional Genomics Research and Crop Improvement
Zhang, H., Zhang, J., Lang, Z., Botella, J. R., & Zhu, J. (2017) · Critical Reviews in Plant Sciences · doi:10.1080/07352689.2017.1402989 · Cited by 179
Efficient CRISPR/Cas9-based genome editing in carrot cells
Klimek‐Chodacka, M., Oleszkiewicz, T., Lowder, L. G., Qi, Y., & Barański, R. (2018) · Plant Cell Reports · doi:10.1007/s00299-018-2252-2 · Cited by 177
PKM1 Confers Metabolic Advantages and Promotes Cell-Autonomous Tumor Cell Growth
Morita, M., Sato, T., Nomura, M., Sakamoto, Y., Inoue, Y., Tanaka, R., Ito, S., Kurosawa, K., Yamaguchi, K., Sugiura, Y., et al. (2018) · Cancer Cell · doi:10.1016/j.ccell.2018.02.004 · Cited by 170
Movement of accessible plasma membrane cholesterol by the GRAMD1 lipid transfer protein complex
Naito, T., Ercan, B., Krshnan, L., Triebl, A., Koh, D. H. Z., Wei, F.-Y., Tomizawa, K., Torta, F. T., Wenk, M. R., & Saheki, Y. (2019) · eLife · doi:10.7554/elife.51401 · Cited by 171
Efficient CRISPR-mediated gene targeting and transgene replacement in the beetleTribolium castaneum
Gilles, A. F., Schinko, J. B., & Averof, M. (2015) · Development · doi:10.1242/dev.125054 · Cited by 175
Saturation variant interpretation using CRISPR prime editing
Erwood, S., Bily, T. M. I., Lequyer, J., Yan, J., Gulati, N., Brewer, R. A., Zhou, L., Pelletier, L., Ivakine, E. A., & Cohn, R. D. (2022) · Nature Biotechnology · doi:10.1038/s41587-021-01201-1 · Cited by 163
Applications of CRISPR/Cas System to Bacterial Metabolic Engineering
Cho, S., Shin, J., & Cho, B. (2018) · International Journal of Molecular Sciences · doi:10.3390/ijms19041089 · Cited by 180
Deneault, E., White, S. H., Rodrigues, D. C., Ross, P. J., Faheem, M., Zaslavsky, K., Wang, Z., Alexandrova, R., Pellecchia, G., Wei, W., et al. (2018) · Stem Cell Reports · doi:10.1016/j.stemcr.2018.10.003 · Cited by 145
Multiplexed precision genome editing with trackable genomic barcodes in yeast
Roy, K. R., Smith, J., Vonesch, S. C., Lin, G., Szu‐Tu, C., Lederer, A. R., Chu, A. M., Suresh, S., Nguyen, M., Horecka, J., et al. (2018) · Nature Biotechnology · doi:10.1038/nbt.4137 · Cited by 175
Ratz, M., Testa, I., Hell, S. W., & Jakobs, S. (2015) · Scientific Reports · doi:10.1038/srep09592 · Cited by 166
Kozono, S., Lin, Y.-M., Seo, H.-S., Pinch, B., Lian, X., Qiu, C., Herbert, M. K., Chen, C.-H., Tan, L., Gao, Z. J., et al. (2018) · Nature Communications · doi:10.1038/s41467-018-05402-2 · Cited by 168
Zebrafish Genome Engineering Using the CRISPR–Cas9 System
Li, M., Zhao, L., Page-McCaw, P., & Chen, W. (2016) · Trends in Genetics · doi:10.1016/j.tig.2016.10.005 · Cited by 181
Engineered dCas9 with reduced toxicity in bacteria: implications for genetic circuit design
Zhang, S., & Voigt, C. A. (2018) · Nucleic Acids Research · doi:10.1093/nar/gky884 · Cited by 171
Metadata
- Authors
- Hsu, P. D., Scott, D. A., Weinstein, J. A., Ran, F. A., Konermann, S., Agarwala, V., Li, Y., Fine, E. J., Wu, X., Shalem, O., Cradick, T. J., Marraffini, L. A., Bao, G., & Zhang, F.
- Affiliations
- Broad Institute (US)
- Georgia Institute of Technology (US)
- Koch Institute for Integrative Cancer Research At MIT (US)
- Rockefeller University (US)
- Corresponding author
- Zhang, Feng
- ORCID
- Venue
- Nature Biotechnology
- Published
- 2013-07-21
- Volume
- 31
- Issue
- 9
- Pages
- 827-832
- ISSN
- 0733-222X (ISSN-L)
- 1087-0156
- 1546-1696
- PMC
- PMC3969858
- OpenAlex
- W2036176224
- Cited by
- 4,741
- References
- 29
- Citations by year
- 2026: 242 · 2025: 300 · 2024: 304 · 2023: 331 · 2022: 372 · 2021: 424 · 2020: 416 · 2019: 422 · 2018: 463 · 2017: 438 · 2016: 402 · 2015: 297 · 2014: 269 · 2013: 53
- FWCI
- 152.56
- Type
- journal article
- Language
- en
- Open access
- green
- europepmc.org
- Licence
- www.springer.com/tdm (tdm)
- Locations
- Nature Biotechnology (publishedVersion)
- PubMed (publishedVersion)
- Europe PMC (PubMed Central) (acceptedVersion)
- PubMed Central (acceptedVersion)
- DSpace@MIT (Massachusetts Institute of Technology) (submittedVersion)
- PMC
- Funders
- National Science Foundation
- Howard Hughes Medical Institute
- Damon Runyon Cancer Research Foundation
- McKnight Foundation
- Life Sciences Research Foundation
- National Institutes of Health (R01‐grant, GM34277)
- Harvard Stem Cell Institute
- National Institute of Mental Health (DP1MH100706)
- National Eye Institute (PN2EY018244)
- National Cancer Institute (R01CA133404)
- National Institute of General Medical Sciences (R01GM034277, R01 GM34277, T32GM007753)
- National Institute of Diabetes and Digestive and Kidney Diseases (R01DK097768)
- National Center for Chronic Disease Prevention and Health Promotion (DP1‑MH100706)
- 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 › RNA Interference and Gene Delivery
- Keywords
- SpCas9, single guide RNA, genome editing, off-target effects, mismatch tolerance, DNA methylation, indel mutations, guide RNA design, target site selection, HEK293T cells, web-based design tool
- MeSH terms
- Bacterial Proteins, Base Sequence, Deoxyribonucleases, DNA, Genetic Engineering, Molecular Sequence Data, Streptococcus pyogenes, Transcription Factors, Base Pair Mismatch, RNA, Small Untranslated
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