The DOI matches its Crossref record; 11 of the 11 references it cites were found in a registry or an index.
Efficient Multiplexed Integration of Synergistic Alleles and Metabolic Pathways in Yeasts via CRISPR-Cas
Open accessAbstract
CRISPR-Cas genome engineering in yeast has relied on preparation of complex expression plasmids for multiplexed gene knockouts and point mutations. Here we show that co-transformation of a single linearized plasmid with multiple PCR-generated guide RNA (gRNA) and donor DNA cassettes facilitates high-efficiency multiplexed integration of point mutations and large constructs. This technique allowed recovery of marker-less triple-engineering events with 64% efficiency without selection for expression of all gRNAs. The gRNA cassettes can be easily made by PCR and delivered in any combination. We employed this method to rapidly phenotype up to five specific allele combinations and identify synergistic effects. To prototype a pathway for the production of muconic acid, we integrated six DNA fragments totaling 24 kb across three loci in naive Saccharomyces cerevisiae in a single transformation. With minor modifications, we integrated a similar pathway in Kluyveromyces lactis. The flexibility afforded by combinatorial gRNA delivery dramatically accelerates complex strain engineering for basic research and industrial fermentation.
Overview
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
Verification
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.
Self-check
Percentages, sums and reported differences re-computed from the paper’s own numbers.
The self-check needs the paper’s full text.
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
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
DNA targeting specificity of RNA-guided Cas9 nucleases
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., et al. (2013) · Nature Biotechnology · doi:10.1038/nbt.2647 · Cited by 4,741
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
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 · cited 1×
“The NGG sequence is referred to as a protospacer adjacent motif (PAM) sequence, and the 8 bp of DNA preceding the PAM sequence are especially important for enforcing specificity (Fu et al., 2013; Hsu et al., 2013).”
BackgroundTargeted genome engineering in human cells with the Cas9 RNA-guided endonuclease
Cho, S. W., Kim, S., Kim, J. M., & Kim, J.-S. (2013) · Nature Biotechnology · doi:10.1038/nbt.2507 · Cited by 1,863
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 · cited 2×
“coli, the yeast equivalent (YOGE) functions at a much lower efficiency (Wang et al., 2009; DiCarlo et al., 2013b).”
BackgroundEngineering Yeast Transcription Machinery for Improved Ethanol Tolerance and Production
Alper, H., Moxley, J., Nevoigt, E., Fink, G. R., & Stephanopoulos, G. (2006) · Science · doi:10.1126/science.1131969 · Cited by 823 · cited 1×
“Finally we tested a series of five alleles associated with resistance to elevated ethanol concentrations (SPT15 F177S, SPT15 Y195H, SPT15 K218R, PRO1 D154N, and PUT1 deletion) (Takagi et al., 2005; Alper et al., 2006).”
Methodvan Dijken, J. P., Bauer, J., Brambilla, L., Duboc, P., Francois, J. M., Gancedo, C., Giuseppin, M. L. F., Heijnen, J. J., Hoare, M., Lange, H. C., et al. (2000) · Enzyme and Microbial Technology · doi:10.1016/s0141-0229(00)00162-9 · Cited by 598 · cited 1×
“PK2-1C (van Dijken et al., 2000) (Figure 1).”
Cited by
A gRNA-tRNA array for CRISPR-Cas9 based rapid multiplexed genome editing in Saccharomyces cerevisiae
Zhang, Y., Wang, J., Wang, Z., Zhang, Y., Shi, S., Nielsen, J., & Liu, Z. (2019) · Nature Communications · doi:10.1038/s41467-019-09005-3 · Cited by 294
Bricks and blueprints: methods and standards for DNA assembly.
Casini, A., Storch, M., Baldwin, G. S., & Ellis, T. (2015) · Nature Reviews Molecular Cell Biology · doi:10.1038/nrm4014 · Cited by 286
Simplified CRISPR-Cas genome editing for Saccharomyces cerevisiae
Generoso, W. C., Gottardi, M., Oreb, M., & Boles, E. (2016) · Journal of Microbiological Methods · doi:10.1016/j.mimet.2016.06.020 · Cited by 165
Pyne, M. E., Gold, N. D., & Martin, V. J. J. (2023) · Metabolic Engineering · doi:10.1016/j.ymben.2023.03.010 · Cited by 37
Metadata
- Authors
- Horwitz, A. A., Walter, J. M., Schubert, M. G., Kung, S. H., Hawkins, K., Platt, D. M., Hernday, A. D., Mahatdejkul-Meadows, T., Szeto, W., Chandran, S. S., & Newman, J. D.
- Venue
- Cell Systems
- Publisher
- Elsevier BV
- Published
- 2015-07
- Volume
- 1
- Issue
- 1
- Pages
- 88-96
- ISSN
- 2405-4712 (ISSN-L)
- OpenAlex
- W2043784202
- Cited by
- 297
- References
- 11
- FWCI
- 14.92
- Type
- journal article
- Language
- en
- Open access
- hybrid
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
How to cite
Cite CoChat for the checks on this page, and the paper for its own findings.
- This page
- CoChat. (2026). Citation check: Efficient Multiplexed Integration of Synergistic Alleles and Metabolic Pathways in Yeasts via CRISPR-Cas. https://app.cochat.ai/papers/10.1016/j.cels.2015.02.001
- The paper
- Horwitz, A. A., Walter, J. M., Schubert, M. G., Kung, S. H., Hawkins, K., Platt, D. M., Hernday, A. D., Mahatdejkul-Meadows, T., Szeto, W., Chandran, S. S., & Newman, J. D. (2015). Efficient Multiplexed Integration of Synergistic Alleles and Metabolic Pathways in Yeasts via CRISPR-Cas. Cell Systems. https://doi.org/10.1016/j.cels.2015.02.001
Data sources
Bibliographic data: Semantic Scholar, OpenAlex, Crossref.
Contains information from the Semantic Scholar Academic Graph Datasets, made available under the ODC Attribution License (ODC-BY 1.0).
Semantic Scholar’s data is described in: Kinney, R., et al. (2023). The Semantic Scholar Open Data Platform. arXiv:2301.10140.
