Multigene editing in the Escherichia coli genome via the CRISPR-Cas9 system
Open accessAbstract
An efficient genome-scale editing tool is required for construction of industrially useful microbes. We describe a targeted, continual multigene editing strategy that was applied to the Escherichia coli genome by using the Streptococcus pyogenes type II CRISPR-Cas9 system to realize a variety of precise genome modifications, including gene deletion and insertion, with a highest efficiency of 100%, which was able to achieve simultaneous multigene editing of up to three targets. The system also demonstrated successful targeted chromosomal deletions in Tatumella citrea, another species of the Enterobacteriaceae, with highest efficiency of 100%.
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?
The type II CRISPR/Cas9 system has been exploited for high-efficiency gene editing in model microorganisms such as E. coli, S. cerevisiae, and Streptococcus pneumoniae.
“The type II CRISPR/Cas9 system has been exploited for high-efficiency gene editing in model microorganisms such as E. coli (Jiang et al., 2015), S. cerevisiae (DiCarlo et al., 2013), and Streptococcus pneumoniae (Jiang et al., 2013).”
From Gas Fermentation—A Flexible Platform for Commercial Scale Production of Low-Carbon-Fuels and Chemicals from Waste and Renewable Feedstocks (2016) · cites this paper as Jiang et al., 2015
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
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 · see citing works
One-step inactivation of chromosomal genes in Escherichia coli K-12 using PCR products.
Datsenko, K., & Wanner, B. (2000) · Proceedings of the National Academy of Sciences of the United States of America · doi:10.1073/pnas.120163297 · Cited by 15,902 · see citing works
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 · see citing works
RNA-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 · see citing works
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 · cited 3× · see citing works
“coli (24, 35), which separated cas9 and the sgRNA in pCas and pTarget series, respectively (Fig.”
BackgroundMethodGuzman, L., Belin, D., Carson, M. J., & Beckwith, J. (1995) · Journal of Bacteriology · doi:10.1128/jb.177.14.4121-4130.1995 · Cited by 4,942 · see citing works
Double nicking by RNA-guided CRISPR Cas9 for enhanced genome editing specificity
Ran, F. A., Hsu, P. D., Lin, C.-Y., Gootenberg, J. S., Konermann, S., Trevino, A. E., Scott, D. A., Inoue, A., Matoba, S., Zhang, Y., et al. (2013) · Cell · doi:10.1016/j.cell.2013.08.021 · Cited by 3,299 · see citing works
High-Frequency off-Target Mutagenesis Induced by CRISPR-Cas Nucleases in Human Cells
Fu, Y., Foden, J. A., Khayter, C., Maeder, M. L., Reyon, D., Joung, J. K., & Sander, J. D. (2013) · Nature Biotechnology · doi:10.1038/nbt.2623 · Cited by 3,292 · see citing works
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 · see citing works
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 · see citing works
Targeted genome modification of crop plants using a CRISPR-Cas system
Shan, Q., Wang, Y., Li, J., Zhang, Y., Chen, K., Liang, Z., Zhang, K., Liu, J., Xi, J., Qiu, J., et al. (2013) · Nature Biotechnology · doi:10.1038/nbt.2650 · Cited by 1,965 · see citing works
Non-fermentative pathways for synthesis of branched-chain higher alcohols as biofuels
Atsumi, S., Hanai, T., & Liao, J. C. (2008) · Nature · doi:10.1038/nature06450 · Cited by 1,945 · see citing works
Genetic applications of an inverse polymerase chain reaction.
Ochman, H., Gerber, A. S., & Hartl, D. L. (1988) · Genetics · doi:10.1093/genetics/120.3.621 · Cited by 1,868 · see citing works
Engineering a mevalonate pathway in Escherichia coli for production of terpenoids
Martin, V. J. J., Pitera, D. J., Withers, S. T., Newman, J. D., & Keasling, J. D. (2003) · Nature Biotechnology · doi:10.1038/nbt833 · Cited by 1,720 · see citing works
An efficient recombination system for chromosome engineering in Escherichia coli
Yu, D., Ellis, H. M., Lee, E.-C., Jenkins, N. A., Copeland, N. G., & Court, D. L. (2000) · Proceedings of the National Academy of Sciences · doi:10.1073/pnas.100127597 · Cited by 1,688 · see citing works
Genome engineering in Saccharomyces cerevisiae using CRISPR-Cas systems
DiCarlo, J. E., Norville, J. E., Mali, P., Rios, X., Aach, J., & Church, G. M. (2013) · Nucleic Acids Research · doi:10.1093/nar/gkt135 · Cited by 1,566 · see citing works
Programming cells by multiplex genome engineering and accelerated evolution
Wang, H. H., Isaacs, F. J., Carr, P. A., Sun, Z. Z., Xu, G., Forest, C. R., & Church, G. M. (2009) · Nature · doi:10.1038/nature08187 · Cited by 1,529 · see citing works
Gust, B., Challis, G. L., Fowler, K., Kieser, T., & Chater, K. F. (2003) · Proceedings of the National Academy of Sciences · doi:10.1073/pnas.0337542100 · Cited by 1,416 · see citing works
Microbial production of fatty-acid-derived fuels and chemicals from plant biomass
Steen, E. J., Kang, Y., Bokinsky, G., Hu, Z., Schirmer, A., McClure, A., del Cardayré, S. B., & Keasling, J. D. (2010) · Nature · doi:10.1038/nature08721 · Cited by 1,338 · see citing works
Structural basis of PAM-dependent target DNA recognition by the Cas9 endonuclease
Anders, C., Niewoehner, O., Duerst, A., & Jínek, M. (2014) · Nature · doi:10.1038/nature13579 · Cited by 1,319 · see citing works
Malard, F., Wolter, A. C., Marquevielle, J., et al. (2024) · HAL (Le Centre pour la Communication Scientifique Directe) · doi:10.1093/nar · Cited by 1,259 · see citing works
A new logic for DNA engineering using recombination in Escherichia coli
Zhang, Y., Buchholz, F., Muyrers, J. P. P., & Stewart, A. F. (1998) · Nature Genetics · doi:10.1038/2417 · Cited by 1,225 · see citing works
Metabolic engineering of Escherichia coli for direct production of 1,4-butanediol
Yim, H. H., Haselbeck, R. J., Niu, W., Pujol-Baxley, C., Burgard, A. P., Boldt, J., Khandurina, J., Trawick, J. D., Osterhout, R. E., Stephen, R., et al. (2011) · Nature Chemical Biology · doi:10.1038/nchembio.580 · Cited by 1,119 · see citing works
Amann, E. P., Ochs, B., & Kj, A. (1988) · Gene · doi:10.1016/0378-1119(88)90440-4 · Cited by 1,045 · see citing works
The Next-Generation Sequencing Revolution and Its Impact on Genomics
Koboldt, D. C., Steinberg, K. M., Larson, D. E., Wilson, R. K., & Mardis, E. R. (2013) · Cell · doi:10.1016/j.cell.2013.09.006 · Cited by 1,039 · see citing works
Metabolic engineering for the microbial production of 1,3-propanediol.
Nakamura, C. E., & Whited, G. (2003) · Current Opinion in Biotechnology · doi:10.1016/j.copbio.2003.08.005 · Cited by 801 · see citing works
Recombineering: a homologous recombination-based method of genetic engineering
Sharan, S. K., Thomason, L. C., Kuznetsov, S. G., & Court, D. L. (2009) · Nature Protocols · doi:10.1038/nprot.2008.227 · Cited by 788 · see citing works
Biotechnological production of amino acids and derivatives: current status and prospects
Leuchtenberger, W., Huthmacher, K., & Drauz, K. (2005) · Applied Microbiology and Biotechnology · doi:10.1007/s00253-005-0155-y · Cited by 777 · see citing works
Engineering BioBrick vectors from BioBrick parts
Shetty, R. P., Endy, D., & Knight, T. F. (2008) · Journal of Biological Engineering · doi:10.1186/1754-1611-2-5 · Cited by 755 · see citing works
CRISPR/Cas9 systems targeting β-globin and CCR5 genes have substantial off-target activity
Cradick, T. J., Fine, E. J., Antico, C. J., & Bao, G. (2013) · Nucleic Acids Research · doi:10.1093/nar/gkt714 · Cited by 590 · see citing works
Cobb, R. E., Wang, Y., & Zhao, H. (2014) · ACS Synthetic Biology · doi:10.1021/sb500351f · Cited by 566 · see citing works
McDaniel, R. N., Thamchaipenet, A., Gustafsson, C., Fu, H., Betlach, M. C., Betlach, M. C., & Ashley, G. W. (1999) · Proceedings of the National Academy of Sciences · doi:10.1073/pnas.96.5.1846 · Cited by 463 · see citing works
Ancestral Resurrection and Directed Evolution of Fungal Mesozoic Laccases
Agans, R., Gordon, A., Kramer, D. L., Pérez-Burillo, S., Rufián‐Henares, J. Á., & Paliy, O. (2018) · Institutional Repository of the University of Granada (University of Granada) · doi:10.1128/aem · Cited by 459 · see citing works
Siegele, D. A., & Hu, J. C. (1997) · Proceedings of the National Academy of Sciences · doi:10.1073/pnas.94.15.8168 · Cited by 418 · see citing works
Highly Efficient Genome Modifications Mediated by CRISPR/Cas9 in Drosophila
Yu, Z., Ren, M., Wang, Z., Zhang, B., Rong, Y. S., Jiao, R., & Gao, G. (2013) · Genetics · doi:10.1534/genetics.113.153825 · Cited by 339 · see citing works
Genome‐scale engineering for systems and synthetic biology
Esvelt, K. M., & Wang, H. H. (2013) · Molecular Systems Biology · doi:10.1038/msb.2012.66 · Cited by 319 · see citing works
Martinez, E., Bartolomé, B., & de la Cruz, F. (1988) · Gene · doi:10.1016/0378-1119(88)90608-7 · Cited by 302 · see citing works
The CRISPR/Cas System mediates efficient genome engineering in Bombyx mori
Wang, Y., Li, Z., Xu, J. Q., Zeng, B., Lin, L., You, L., Chen, Y., Huang, Y., & Tan, A. (2013) · Cell Research · doi:10.1038/cr.2013.146 · Cited by 299 · see citing works
Metabolic Engineering for Microbial Production of Aromatic Amino Acids and Derived Compounds
Bongaerts, J., Krämer, M., Müller, U., Raeven, L. J. R. M., & Wubbolts, M. G. (2001) · Metabolic Engineering · doi:10.1006/mben.2001.0196 · Cited by 290 · see citing works
Rapid profiling of a microbial genome using mixtures of barcoded oligonucleotides
Warner, J. R., Reeder, P. J., Karimpour‐Fard, A., Woodruff, L. B. A., & Gill, R. T. (2010) · Nature Biotechnology · doi:10.1038/nbt.1653 · Cited by 284 · see citing works
Enhanced levels of λ Red-mediated recombinants in mismatch repair mutants
Costantino, N., & Court, D. L. (2003) · Proceedings of the National Academy of Sciences · doi:10.1073/pnas.2434959100 · Cited by 275 · see citing works
Pósfai, G., Kolisnychenko, V., Bereczki, Z., & Blattner, F. R. (1999) · Nucleic Acids Research · doi:10.1093/nar/27.22.4409 · Cited by 252 · see citing works
Group II introns as controllable gene targeting vectors for genetic manipulation of bacteria
Karberg, M., Guo, H., Zhong, J., Coon, R., Perutka, J., & Lambowitz, A. M. (2001) · Nature Biotechnology · doi:10.1038/nbt1201-1162 · Cited by 222 · see citing works
An end-joining repair mechanism in Escherichia coli
Chayot, R., Montagne, B., Mazel, D., et al. (2010) · Proceedings of the National Academy of Sciences · doi:10.1073/pnas.0906355107 · Cited by 171 · see citing works
Nupan, B., Phongdara, A., Saengsakda, M., et al. (2010) · Developmental & Comparative Immunology · doi:10.1016/j.dci.2010.11.016 · Cited by 136 · see citing works
Multiplexed Genome Engineering and Genotyping Methods
Wang, H. H., & Church, G. M. (2011) · Methods in enzymology on CD-ROM/Methods in enzymology · doi:10.1016/b978-0-12-385120-8.00018-8 · Cited by 132 · see citing works
(2019) · Protocol Exchange · doi:10.1038/nprot · Cited by 111 · see citing works
Multiple mechanisms controlling carbon metabolism in bacteria
Saier, M. H. (1998) · Biotechnology and Bioengineering · doi:10.1002/(sici)1097-0290(19980420)58:2/3 · Cited by 97 · see citing works
Rapid and efficient construction of markerless deletions in the Escherichia coli genome
Yu, B., Kang, K. H., Lee, Y., Sung, B. H., Kim, M. S., & Kim, S. (2008) · Nucleic Acids Research · doi:10.1093/nar/gkn359 · Cited by 96 · see citing works
Yang, J., Sun, B., Huang, H., Jiang, Y., Diao, L., Chen, B., Xu, C., Wang, X., Liu, J., Jiang, W., et al. (2014) · Applied and Environmental Microbiology · doi:10.1128/aem.00313-14 · Cited by 87 · see citing works
Self-citationGeneralized bacterial genome editing using mobile group II introns and Cre‐lox
Enyeart, P. J., Chirieleison, S. M., Dao, M. N., Perůtka, J., Quandt, E. M., Yao, J., Whitt, J. T., Keatinge‐Clay, A. T., Lambowitz, A. M., & Ellington, A. D. (2013) · Molecular Systems Biology · doi:10.1038/msb.2013.41 · Cited by 82 · see citing works
Non-homologous end-joining: bacteria join the chromosome breakdance
Wilson, T. E., Topper, L. M., & Palmbos, P. L. (2003) · Trends in Biochemical Sciences · doi:10.1016/s0968-0004(03)00005-7 · Cited by 74 · see citing works
Zhang, Q., Rho, M., Tang, H., Doak, T. G., & Ye, Y. (2013) · Genome biology · doi:10.1186/gb-2013-14-4-r40 · Cited by 59 · see citing works
Pujol, C. J., & Kado, C. I. (2000) · Journal of Bacteriology · doi:10.1128/jb.182.8.2230-2237.2000 · Cited by 51 · see citing works
Malyarchuk, S., Wright, D., Castore, R., et al. (2007) · DNA repair · doi:10.1016/j.dnarep.2007.04.004 · Cited by 46 · see citing works
Cha, J. S., Pujol, C. J., & Kado, C. I. (1997) · Applied and Environmental Microbiology · doi:10.1128/aem.63.1.71-76.1997 · Cited by 34 · see citing works
Banta, S. A., Boston, M., Jarnagin, A., et al. (2002) · Metabolic Engineering · doi:10.1006/mben.2002.0231 · Cited by 16 · see citing works
Flagged first, then by citations
Research works printed but not linked: 1
- [1]1. Ingram LO, Gomez PF, Lai X, Moniruzzaman M, Wood BE, Yomano LP, York SW. 1998. Metabolic engineering of bacteria for ethanol production. Biotechnol Bioeng 58: 204-214. http://dx.doi.org/10.1002 /(SICI)1097-0290(19980420)58:2/3 < 204::AID-BIT13 > 3.0.CO;2-C.
Cited by
Sorted by citations
Liew, F., Martin, M. E., Tappel, R. C., Heijstra, B. D., Mihalcea, C., & Köpke, M. (2016) · Frontiers in Microbiology · doi:10.3389/fmicb.2016.00694 · Cited by 485 · see citing works
Cpf1 Is a Versatile Tool for CRISPR Genome Editing Across Diverse Species of Cyanobacteria
Ungerer, J., & Pakrasi, H. B. (2016) · Scientific Reports · doi:10.1038/srep39681 · Cited by 308 · see citing works
Toolboxes for cyanobacteria: recent advances and future direction
Sun, T., Li, S., Song, X., Diao, J., Chen, L., & Zhang, W. (2018) · Biotechnology Advances · doi:10.1016/j.biotechadv.2018.04.007 · Cited by 116 · see citing works
Characterization and Repurposing of Type I and Type II CRISPR–Cas Systems in Bacteria
Hidalgo-Cantabrana, C., Goh, Y. J., & Barrangou, R. (2019) · Journal of Molecular Biology · doi:10.1016/j.jmb.2018.09.013 · Cited by 63 · see citing works
Modulating co-translational protein folding by rational design and ribosome engineering
Ahn, M., Włodarski, T., Mitropoulou, A., Chan, S. H. S., Sidhu, H., Plessa, E., Becker, T. A., Budisa, N., Waudby, C. A., Beckmann, R., et al. (2022) · Nature Communications · doi:10.1038/s41467-022-31906-z · Cited by 45 · see citing works
Potential Role of Lysine Acetylation in Antibiotic Resistance of Escherichia coli
Fang, Z., Lai, F., Cao, K., Zhang, Z., Cao, L., Liu, S., Duan, Y., Yin, X., Ge, R., He, Q.-Y., et al. (2022) · mSystems · doi:10.1128/msystems.00649-22 · Cited by 40 · see citing works
Meng, F., Liu, Y., Nie, T., Tang, C., Lyu, F., Bie, X., Lu, Y., Zhao, M., & Lu, Z. (2022) · Applied and Environmental Microbiology · doi:10.1128/aem.00371-22 · Cited by 24 · see citing works
Development of a base editor for convenient and multiplex genome editing in cyanobacteria
Li, X.-D., Liu, L.-M., Xi, Y.-C., Sun, Q.-W., Luo, Z., Huang, H.-L., Wang, X.-W., Jiang, H.-B., & Chen, W. (2024) · Communications Biology · doi:10.1038/s42003-024-06696-3 · Cited by 14 · see citing works
- Authors
- Jiang, Y., Chen, B., Duan, C., Sun, B., Yang, J., & Yang, S.
- Affiliations
- Chinese Academy of Sciences (CN)
- Venue
- Applied and Environmental Microbiology
- Publisher
- American Society for Microbiology
- Published
- 2015-04
- Volume
- 81
- Issue
- 7
- Pages
- 2506-2514
- ISSN
- 0099-2240 (ISSN-L)
- 1098-5336
- PMC
- PMC4357945
- OpenAlex
- W2120263648
- Cited by
- 1,362
- References
- 57
- Citations by year
- 2026: 134 · 2025: 188 · 2024: 183 · 2023: 160 · 2022: 142 · 2021: 130 · 2020: 135 · 2019: 86 · 2018: 87 · 2017: 49 · 2016: 45 · 2015: 22
- FWCI
- 29.37
- Type
- journal article
- Language
- en
- Update notices
- correction (2016-05-31)
- Open access
- bronze
- doi.org
- Locations
- Applied and Environmental Microbiology (publishedVersion)
- PubMed Central (publishedVersion)
- PubMed (publishedVersion)
- Topics
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › CRISPR and Genetic Engineering
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Microbial Metabolic Engineering and Bioproduction
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Genetics › Bacterial Genetics and Biotechnology
- Keywords
- CRISPR/Cas9, Streptococcus pyogenes, Escherichia coli genome, targeted gene deletion, gene insertion, Enterobacteriaceae
- MeSH terms
- Enterobacteriaceae, Genes, Bacterial, Genetics, Microbial, Molecular Biology, Recombination, Genetic, Streptococcus pyogenes, Genome, Bacterial, CRISPR-Cas Systems
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
