RNA targeting with CRISPR–Cas13
Open accessAbstract
RNA has important and diverse roles in biology, but molecular tools to manipulate and measure it are limited. For example, RNA interference can efficiently knockdown RNAs, but it is prone to off-target effects, and visualizing RNAs typically relies on the introduction of exogenous tags. Here we demonstrate that the class 2 type VI RNA-guided RNA-targeting CRISPR-Cas effector Cas13a (previously known as C2c2) can be engineered for mammalian cell RNA knockdown and binding. After initial screening of 15 orthologues, we identified Cas13a from Leptotrichia wadei (LwaCas13a) as the most effective in an interference assay in Escherichia coli. LwaCas13a can be heterologously expressed in mammalian and plant cells for targeted knockdown of either reporter or endogenous transcripts with comparable levels of knockdown as RNA interference and improved specificity. Catalytically inactive LwaCas13a maintains targeted RNA binding activity, which we leveraged for programmable tracking of transcripts in live cells. Our results establish CRISPR-Cas13a as a flexible platform for studying RNA in mammalian cells and therapeutic development.
The claims and statistics of this paper haven’t been checked yet.
Run the check — free, no account needed. It reads the open-access text and checks the sources it cites.
Integrity signals
Registry
Cites
Statistics
Claims the paper makes
The claims and statistics of this paper haven’t been checked yet.
Run the check — free, no account needed. It reads the open-access text and checks the sources it cites.
Claims other papers make about this one
Each is checked against this paper’s own text: does it say what the citing paper says it does?
Structural studies including those using diffusion tensor imaging (DTI) and functional MRI (fMRI) have accelerated understanding of how altered neural circuits relate to clinical symptoms of ASD.
“structural studies including those using diffusion tensor imaging (DTI) 62 and functional MRI (fMRI) 63 have accelerated our understanding of how altered neural circuits relate to clinical symptoms of ASD 64,65”
From Autism spectrum disorder (2020) · cites this paper as [64]
The Extended Data and Methods passages describe Cas13-based RNA imaging in cells, not neural circuits or clinical symptoms of ASD. Searches for ASD, neural circuits, DTI, and fMRI found no passage addressing the attributed claim; the indexed copy is not marked as truncated. (LLM, conf=0.78)
Experimental — accuracy not yet measured
Self-check
Percentages, sums and reported differences re-computed from the paper’s own numbers.
The self-check needs the paper’s full text.
Cites
1 retracted
RETRACTED: The RasGAP-associated endoribonuclease G3BP assembles stress granules
Tourrière, H., Chébli, K., Zekri, L., Courselaud, B., Blanchard, J. M., Bertrand, É., & Tazi, J. (2003) · The Journal of Cell Biology · doi:10.1083/jcb.200212128 · Cited by 834 · see citing works
RetractedFiji: an open-source platform for biological-image analysis
Schindelin, J., Arganda-Carreras, I., Frise, E., Kaynig, V., Longair, M., Pietzsch, T., Preibisch, S., Rueden, C., Saalfeld, S., Schmid, B., et al. (2012) · Nature Methods · doi:10.1038/nmeth.2019 · Cited by 66,890 · see citing works
Subramanian, A., Tamayo, P., Mootha, V. K., Mukherjee, S., Ebert, B. L., Gillette, M. A., Paulovich, A., Pomeroy, S. L., Golub, T. R., Lander, E. S., et al. (2005) · Proceedings of the National Academy of Sciences · doi:10.1073/pnas.0506580102 · Cited by 53,625 · see citing works
Self-citationRSEM: accurate transcript quantification from RNA-Seq data with or without a reference genome
Li, B., & Dewey, C. N. (2011) · BMC Bioinformatics · doi:10.1186/1471-2105-12-323 · Cited by 22,179 · see citing works
Potent and specific genetic interference by double-stranded RNA in Caenorhabditis elegans
Fire, A., Xu, S., Montgomery, M. K., Kostas, S. A., Driver, S. E., & Mello, C. C. (1998) · Nature · doi:10.1038/35888 · Cited by 15,429 · see citing works
Duplexes of 21-nucleotide RNAs mediate RNA interference in cultured mammalian cells
Elbashir, S. M., Harborth, J., Lendeckel, W., Yalcin, A., Weber, K., & Tuschl, T. (2001) · Nature · doi:10.1038/35078107 · Cited by 9,456 · see citing works
Nucleic acid detection with CRISPR-Cas13a/C2c2
Gootenberg, J. S., Abudayyeh, O. O., Lee, J. W., Essletzbichler, P., Dy, A. J., Joung, J., Verdine, V. K., Donghia, N. M., Daringer, N. M., Freije, C. A., et al. (2017) · Science · doi:10.1126/science.aam9321 · Cited by 3,887 · see citing works
Self-citationIn vivo genome editing using Staphylococcus aureus Cas9
Ran, F. A., Cong, L., Yan, W. X., Scott, D. A., Gootenberg, J. S., Kriz, A. J., Zetsche, B., Shalem, O., Wu, X., Makarova, K. S., et al. (2015) · Nature · doi:10.1038/nature14299 · Cited by 2,687 · see citing works
Self-citationExpression profiling reveals off-target gene regulation by RNAi
Jackson, A. L., Bartz, S. R., Schelter, J., Kobayashi, S. V., Burchard, J., Mao, M., Li, B., Cavet, G., & Linsley, P. S. (2003) · Nature Biotechnology · doi:10.1038/nbt831 · Cited by 2,370 · see citing works
C2c2 is a single-component programmable RNA-guided RNA-targeting CRISPR effector
Abudayyeh, O. O., Gootenberg, J. S., Konermann, S., Joung, J., Slaymaker, I. M., Cox, D. B. T., Shmakov, S., Makarova, K. S., Semenova, E., Станиславович, М. Л., et al. (2016) · Science · doi:10.1126/science.aaf5573 · Cited by 2,328 · see citing works
Self-citationPrinciples and Properties of Stress Granules
Protter, D. S. W., & Parker, R. (2016) · Trends in Cell Biology · doi:10.1016/j.tcb.2016.05.004 · Cited by 1,738 · see citing works
The highways and byways of mRNA decay
Garneau, N. L., Wilusz, J., & Wilusz, C. J. (2007) · Nature Reviews Molecular Cell Biology · doi:10.1038/nrm2104 · Cited by 1,294 · see citing works
Genome-scale CRISPR-Cas9 Knockout and Transcriptional Activation Screening
Joung, J., Konermann, S., Gootenberg, J. S., Abudayyeh, O. O., Platt, R. J., Brigham, M. D., Sanjana, N. E., & Zhang, F. (2017) · Nature Protocols · doi:10.1038/nprot.2017.016 · Cited by 1,243 · see citing works
Self-citationDiscovery and Functional Characterization of Diverse Class 2 CRISPR-Cas Systems
Shmakov, S., Abudayyeh, O. O., Makarova, K. S., Wolf, Y. I., Gootenberg, J. S., Semenova, E., Станиславович, М. Л., Joung, J., Konermann, S., Severinov, K., et al. (2015) · Molecular Cell · doi:10.1016/j.molcel.2015.10.008 · Cited by 1,221 · see citing works
Self-citationJain, M., Nijhawan, A., Tyagi, A. K., & Khurana, J. P. (2006) · Biochemical and Biophysical Research Communications · doi:10.1016/j.bbrc.2006.04.140 · Cited by 1,186 · see citing works
Two distinct RNase activities of CRISPR-C2c2 enable guide-RNA processing and RNA detection
East-Seletsky, A., O’Connell, M. R., Knight, S. C., Burstein, D. E., Cate, J. H. D., Tjian, R., & Doudna, J. A. (2016) · Nature · doi:10.1038/nature19802 · Cited by 1,180 · see citing works
Ageing, neurodegeneration and brain rejuvenation
Wyss-Coray, T. (2016) · Nature · doi:10.1038/nature20411 · Cited by 1,147 · see citing works
Diversity and evolution of class 2 CRISPR–Cas systems
Shmakov, S., Smargon, A. A., Scott, D. A., Cox, D. B. T., Pyzocha, N., Yan, W. X., Abudayyeh, O. O., Gootenberg, J. S., Makarova, K. S., Wolf, Y. I., et al. (2017) · Nature Reviews Microbiology · doi:10.1038/nrmicro.2016.184 · Cited by 1,056 · see citing works
Self-citationZhang, Y., Su, J., Duan, S., Ao, Y., Dai, J., Liu, J., Wang, P., Li, Y., Liu, B., Feng, D., et al. (2011) · Plant Methods · doi:10.1186/1746-4811-7-30 · Cited by 1,010 · see citing works
Hutchinson, J. N., Ensminger, A. W., Clemson, C., Lynch, C. R., Lawrence, J. B., & Chess, A. J. (2007) · BMC Genomics · doi:10.1186/1471-2164-8-39 · Cited by 997 · see citing works
Multiplex gene editing by CRISPR–Cpf1 using a single crRNA array
Zetsche, B., Heidenreich, M., Mohanraju, P., Fedorova, I., Kneppers, J., DeGennaro, E. M., Winblad, N., Choudhury, S. R., Abudayyeh, O. O., Gootenberg, J. S., et al. (2017) · Nature Biotechnology · doi:10.1038/nbt.3737 · Cited by 926 · see citing works
Self-citationRole of Stress Granules and RNA-Binding Proteins in Neurodegeneration: A Mini-Review
Vanderweyde, T. E., Youmans, K., Liu‐Yesucevitz, L., & Wolozin, B. L. (2013) · Gerontology · doi:10.1159/000354170 · Cited by 806 · see citing works
Programmable RNA Tracking in Live Cells with CRISPR/Cas9
Nelles, D. A., Fang, M. Y., O’Connell, M. R., Xu, J. L., Markmiller, S., Doudna, J. A., & Yeo, G. W.-M. (2016) · Cell · doi:10.1016/j.cell.2016.02.054 · Cited by 526 · see citing works
Imaging intracellular RNA distribution and dynamics in living cells
Tyagi, S. (2009) · Nature Methods · doi:10.1038/nmeth.1321 · Cited by 400 · see citing works
Genome-scale loss-of-function screening with a lentiviral RNAi library
Root, D. E., Hacohen, N., Hahn, W. C., Lander, E. S., & Sabatini, D. M. (2006) · Nature Methods · doi:10.1038/nmeth924 · Cited by 358 · see citing works
Self-citationRecombinant Probes for Visualizing Endogenous Synaptic Proteins in Living Neurons
Gross, G. G., Junge, J. A., Mora, R. J., Kwon, H., Olson, C. A., Takahashi, T. T., Liman, E. R., Ellis‐Davies, G. C. R., McGee, A. W., Sabatini, B. L., et al. (2013) · Neuron · doi:10.1016/j.neuron.2013.04.017 · Cited by 357 · see citing works
The impact of target site accessibility on the design of effective siRNAs
Tafer, H., Ameres, S. L., Obernosterer, G., et al. (2008) · Nature Biotechnology · doi:10.1038/nbt1404 · Cited by 309 · see citing works
Orthogonal gene knockout and activation with a catalytically active Cas9 nuclease
Dahlman, J. E., Abudayyeh, O. O., Joung, J., Gootenberg, J. S., Zhang, F., & Konermann, S. (2015) · Nature Biotechnology · doi:10.1038/nbt.3390 · Cited by 266 · see citing works
Self-citationLocal RNA base pairing probabilities in large sequences
Bernhart, S. H., Hofacker, I. L., & Stadler, P. F. (2005) · Bioinformatics · doi:10.1093/bioinformatics/btk014 · Cited by 230 · see citing works
Mann, D. G. J., LaFayette, P. R., Abercrombie, L. L., et al. (2011) · Plant Biotechnology Journal · doi:10.1111/j.1467-7652.2011.00658.x · Cited by 167 · see citing works
Unworth, H., Raguz, S., Edwards, H. J., et al. (2010) · The FASEB Journal · doi:10.1096/fj.09-151142 · Cited by 67 · see citing works
Rath, S. P., Donovan, J., Whitney, G., et al. (2015) · Proceedings of the National Academy of Sciences · doi:10.1073/pnas.1513034112 · Cited by 51 · see citing works
Flagged first, then by citations
Research works printed but not linked: 1
- [10]Gootenberg JS Nucleic acid detection with CRISPR-Cas13a/C2c2 Science In press 2017
Cited by
Sorted by citations
Gene regulation by long non-coding RNAs and its biological functions
Statello, L., Guo, C.-J., Chen, L.-L., & Huarte, M. (2021) · Nature Reviews Molecular Cell Biology · doi:10.1038/s41580-020-00315-9 · Cited by 4,991 · see citing works
The biogenesis, biology and characterization of circular RNAs
Kristensen, L. S., Andersen, M. S., Stagsted, L. V. W., Ebbesen, K. K., Hansen, T. B., & Kjems, J. (2019) · Nature Reviews Genetics · doi:10.1038/s41576-019-0158-7 · Cited by 4,823 · see citing works
Adeno-associated virus vector as a platform for gene therapy delivery
Wang, D., Tai, P. W. L., & Gao, G. (2019) · Nature Reviews Drug Discovery · doi:10.1038/s41573-019-0012-9 · Cited by 2,444 · see citing works
Lord, C., Brugha, T. S., Charman, T., Cusack, J., Dumas, G., Frazier, T., Jones, E. J. H., Jones, R. M., Pickles, A., State, M. W., et al. (2020) · Nature Reviews Disease Primers · doi:10.1038/s41572-019-0138-4 · Cited by 1,859 · see citing works
RNA virus interference via CRISPR/Cas13a system in plants
Aman, R., Ali, Z., Butt, H., Mahas, A., Aljedaani, F., Khan, M. Z., Ding, S., & Mahfouz, M. (2018) · Genome Biology · doi:10.1186/s13059-017-1381-1 · Cited by 1,497 · see citing works
CRISPR-Cas guides the future of genetic engineering
Knott, G. J., & Doudna, J. A. (2018) · Science · doi:10.1126/science.aat5011 · Cited by 1,410 · see citing works
Transcriptome Engineering with RNA-Targeting Type VI-D CRISPR Effectors
Konermann, S., Lotfy, P., Brideau, N. J., Oki, J., Shokhirev, M. N., & Hsu, P. D. (2018) · Cell · doi:10.1016/j.cell.2018.02.033 · Cited by 1,164 · see citing works
Delivering the Messenger: Advances in Technologies for Therapeutic mRNA Delivery
Kowalski, P. S., Rudra, A., Miao, L., & Anderson, D. G. (2019) · Molecular Therapy · doi:10.1016/j.ymthe.2019.02.012 · Cited by 1,127 · see citing works
Applications of CRISPR–Cas in agriculture and plant biotechnology
Zhu, H., Li, C., & Gao, C. (2020) · Nature Reviews Molecular Cell Biology · doi:10.1038/s41580-020-00288-9 · Cited by 870 · see citing works
CRISPR/Cas system: recent advances and future prospects for genome editing
Manghwar, H., Lindsey, K., Zhang, X., & Jin, S. (2019) · Trends in Plant Science · doi:10.1016/j.tplants.2019.09.006 · Cited by 529 · see citing works
“The Cas13 system has high RNA target specificity and efficiency [6].”
BackgroundMassively parallel Cas13 screens reveal principles for guide RNA design
Wessels, H.-H., Méndez-Mancilla, A., Guo, X., Legut, M., Daniloski, Z., & Sanjana, N. E. (2020) · Nature Biotechnology · doi:10.1038/s41587-020-0456-9 · Cited by 418 · see citing works
Increasing the specificity of CRISPR systems with engineered RNA secondary structures
Kocak, D. D., Josephs, E. A., Bhandarkar, V., Adkar, S. S., Kwon, J. B., & Gersbach, C. A. (2019) · Nature Biotechnology · doi:10.1038/s41587-019-0095-1 · Cited by 351 · see citing works
A new era in functional genomics screens
Przybyla, L., & Gilbert, L. A. (2022) · Nature Reviews Genetics · doi:10.1038/s41576-021-00409-w · Cited by 281 · see citing works
Structural Basis for the RNA-Guided Ribonuclease Activity of CRISPR-Cas13d
Zhang, C., Konermann, S., Brideau, N. J., Lotfy, P., Wu, X., Novick, S. J., Strutzenberg, T., Griffin, P. R., Hsu, P. D., & Lyumkis, D. (2018) · Cell · doi:10.1016/j.cell.2018.09.001 · Cited by 276 · see citing works
Programmable RNA targeting with the single-protein CRISPR effector Cas7-11
Özcan, A., Krajeski, R., Ioannidi, E., Lee, B., Gardner, A., Makarova, K. S., Koonin, E. V., Abudayyeh, O. O., & Gootenberg, J. S. (2021) · Nature · doi:10.1038/s41586-021-03886-5 · Cited by 263 · see citing works
Self-citationHigh-fidelity Cas13 variants for targeted RNA degradation with minimal collateral effects
Tong, H., Huang, J., Xiao, Q., He, B., Dong, X., Liu, Y., Yang, X., Han, D., Wang, Z., Wang, X., et al. (2023) · Nature Biotechnology · doi:10.1038/s41587-022-01419-7 · Cited by 230 · see citing works
CRISPR-Based Technologies: Impact of RNA-Targeting Systems
Terns, M. P. (2018) · Molecular Cell · doi:10.1016/j.molcel.2018.09.018 · Cited by 168 · see citing works
RNA-Targeting CRISPR-Cas systems and their applications
Burmistrz, M., Krakowski, K., & Krawczyk-Balska, A. (2020) · International Journal of Molecular Sciences · doi:10.3390/ijms21031122 · Cited by 125 · see citing works
Transcriptome-wide Cas13 guide RNA design for model organisms and viral RNA pathogens
Guo, X., Rahman, J. A., Wessels, H.-H., Méndez-Mancilla, A., Haro, D., Chen, X., & Sanjana, N. E. (2021) · Cell Genomics · doi:10.1016/j.xgen.2021.100001 · Cited by 79 · see citing works
Molecular Therapies for Inherited Retinal Diseases—Current Standing, Opportunities and Challenges
Vázquez-Domínguez, I., Garanto, A., & Collin, R. W. J. (2019) · Genes · doi:10.3390/genes10090654 · Cited by 80 · see citing works
Gene Therapy Intervention in Neovascular Eye Disease: A Recent Update
Lin, F.-L., Wang, P.-Y., Chuang, Y.-F., Wang, J.-H., Wong, V. H. Y., Bui, B. V., & Liu, G.-S. (2020) · Molecular Therapy · doi:10.1016/j.ymthe.2020.06.029 · Cited by 74 · see citing works
Chemically modified guide RNAs enhance CRISPR-Cas13 knockdown in human cells
Méndez-Mancilla, A., Wessels, H.-H., Legut, M., Kadina, A., Mabuchi, M., Walker, J., Robb, G. B., Holden, K., & Sanjana, N. E. (2022) · Cell Chemical Biology · doi:10.1016/j.chembiol.2021.07.011 · Cited by 68 · see citing works
Kelley, C. P., Haerle, M. C., & Wang, E. T. (2022) · Cell Reports · doi:10.1016/j.celrep.2022.111226 · Cited by 61 · see citing works
Emerging Species and Genome Editing Tools: Future Prospects in Cyanobacterial Synthetic Biology
Gale, G. A. R., Schiavon Osorio, A. A., Mills, L. A., Wang, B., Lea-Smith, D. J., & McCormick, A. J. (2019) · Microorganisms · doi:10.3390/microorganisms7100409 · Cited by 61 · see citing works
Lohia, A., Sahel, D. K., Salman, M., Singh, V., Mariappan, I., Mittal, A., & Chitkara, D. (2022) · Asian Journal of Pharmaceutical Sciences · doi:10.1016/j.ajps.2022.02.001 · Cited by 33 · see citing works
Intrinsic targeting of host RNA by Cas13 constrains its utility
Li, Z., Li, Z., Cheng, X., Wang, S., Wang, X., Ma, S., Lu, Z., Zhang, H., Zhao, W., Chen, Z., et al. (2023) · Nature Biomedical Engineering · doi:10.1038/s41551-023-01109-y · Cited by 31 · see citing works
Sharma, K. K., Palakolanu, S. R., Bhattacharya, J., Shankhapal, A. R., & Bhatnagar-Mathur, P. (2022) · Frontiers in Genetics · doi:10.3389/fgene.2022.999207 · Cited by 24 · see citing works
Methods for in vitro CRISPR/CasRx-Mediated RNA Editing
Chuang, Y.-F., Wang, P.-Y., Kumar, S., Lama, S., Lin, F.-L., & Liu, G.-S. (2021) · Frontiers in Cell and Developmental Biology · doi:10.3389/fcell.2021.667879 · Cited by 21 · see citing works
Deep learning and CRISPR-Cas13d ortholog discovery for optimized RNA targeting
Wei, J., Lotfy, P., Faizi, K., Baungaard, S., Gibson, E., Wang, E., Slabodkin, H., Kinnaman, E., Chandrasekaran, S., Kitano, H., et al. (2021) · doi:10.1101/2021.09.14.460134 · Cited by 67 · see citing works
Optimization of specific RNA knockdown in mammalian cells with CRISPR-Cas13
Burris, B. J. D., Molina Vargas, A. M., Park, B. J., & O'Connell, M. R. (2022) · Methods · doi:10.1016/j.ymeth.2022.08.007 · Cited by 13 · see citing works
- Authors
- Abudayyeh, O. O., Gootenberg, J. S., Essletzbichler, P., Han, S., Joung, J., Belanto, J. J., Verdine, V., Cox, D. B. T., Kellner, M. J., Regev, A., Lander, E. S., Voytas, D. F., Ting, A. Y., & Zhang, F.
- Affiliations
- Broad Institute (US)
- Stanford University (US)
- University of Minnesota (US)
- Corresponding author
- Zhang, Feng
- ORCID
- Venue
- Nature
- Published
- 2017-10
- Volume
- 550
- Issue
- 7675
- Pages
- 280-284
- ISSN
- 0028-0836 (ISSN-L)
- 1476-4687
- PMC
- PMC5706658
- OpenAlex
- W2763042654
- Cited by
- 2,093
- References
- 32
- Citations by year
- 2026: 202 · 2025: 255 · 2024: 262 · 2023: 244 · 2022: 298 · 2021: 282 · 2020: 232 · 2019: 177 · 2018: 130 · 2017: 10
- FWCI
- 61.29
- Type
- journal article
- Language
- en
- Open access
- bronze
- doi.org
- Licence
- www.springer.com/tdm (tdm)
- Locations
- Nature (publishedVersion)
- PubMed (publishedVersion)
- PubMed Central (acceptedVersion)
- RePEc: Research Papers in Economics (submittedVersion)
- Funders
- National Science Foundation (FA9550-14-1-0060)
- Howard Hughes Medical Institute
- National Institutes of Health (5r01hg009761-06, 5rm1hg006193-10)
- National Institute of Mental Health (DP1MH100706, 5DP1-MH100706, R01MH110049)
- National Human Genome Research Institute (R01-HG009761, RM1‐HG006193)
- National Cancer Institute (F30CA210382)
- National Institute of General Medical Sciences (T32GM007753)
- National Institute of Allergy and Infectious Diseases (R01AI117043)
- 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
- Cas13a, CRISPR-Cas13, RNA knockdown, RNA binding, live-cell RNA imaging, plant cells, off-target specificity, RNA therapeutics
- MeSH terms
- Gene Editing, Cell Survival, Escherichia coli, Humans, RNA, Stress, Physiological, Substrate Specificity, Genes, Reporter, RNA Interference, Leptotrichia, Cell Line, Tumor, Biocatalysis, Gene Knockdown Techniques, HEK293 Cells, Plant Cells, CRISPR-Cas Systems, CRISPR-Associated Proteins
About these checks
Registry, retraction, Cites and statistics results are computed, not judged, with one exception: a printed reference that no index lists is read once by a model before it counts as a citation error. Claims are extracted by a model and kept only when their quote appears verbatim in the text; each cited source is then checked, with a model’s help, for whether it says what the claim relies on. Those verdicts can be wrong — read the claim’s quote and the cited source before relying on one.
Statistics and arithmetic are recomputed, not judged.
Confidence bands are shown only for fields calibrated on our annotated test set. None is calibrated yet, so no band is shown.
What counts as a claim ›How we measure confidence ›How we check this ›
Are you an author, or do you see a problem with this page? Request a review
