The DOI matches its Crossref record; 6 of the 6 references it cites were found in a registry or an index.
Rapid and Precise Engineering of the Caenorhabditis elegans Genome with Lethal Mutation Co-Conversion and Inactivation of NHEJ Repair
Open accessAbstract
As in other organisms, CRISPR/Cas9 methods provide a powerful approach for genome editing in the nematode Caenorhabditis elegans. Oligonucleotides are excellent repair templates for introducing substitutions and short insertions, as they are cost effective, require no cloning, and appear in other organisms to target changes by homologous recombination at DNA double-strand breaks (DSBs). Here, I describe a methodology in C. elegans to efficiently knock in epitope tags in 8–9 days, using a temperature-sensitive lethal mutation in the pha-1 gene as a co-conversion marker. I demonstrate that 60mer oligos with 29 bp of homology drive efficient knock-in of point mutations, and that disabling nonhomologous end joining by RNAi inactivation of the cku-80 gene significantly improves knock-in efficiency. Homology arms of 35–80 bp are sufficient for efficient editing and DSBs up to 54 bp away from the insertion site produced knock-ins. These findings will likely be applicable for a range of genome editing approaches in C. elegans, which will improve editing efficiency and minimize screening efforts.
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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
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
Efficient 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
Genome-Scale CRISPR-Mediated Control of Gene Repression and Activation
Gilbert, L. A., Horlbeck, M. A., Adamson, B., Villalta, J. E., Chen, Y., Whitehead, E. H., Guimaraes, C., Panning, B., Ploegh, H. L., Bassik, M. C., et al. (2014) · Cell · doi:10.1016/j.cell.2014.09.029 · Cited by 2,728
Cas9 as a versatile tool for engineering biology
Mali, P., Esvelt, K. M., & Church, G. M. (2013) · Nature Methods · doi:10.1038/nmeth.2649 · Cited by 1,197
Heritable genome editing in C. elegans via a CRISPR-Cas9 system
Friedland, A. E., Tzur, Y. B., Esvelt, K. M., Colaiácovo, M. P., Church, G. M., & Calarco, J. A. (2013) · Nature Methods · doi:10.1038/nmeth.2532 · Cited by 899
Cited by
Paix, A., Folkmann, A., Rasoloson, D., & Seydoux, G. (2015) · Genetics · doi:10.1534/genetics.115.179382 · Cited by 675
Zhang, L., Ward, J. D., Cheng, Z., & Dernburg, A. F. (2015) · Development · doi:10.1242/dev.129635 · Cited by 599
Self-citationStreamlined Genome Engineering with a Self-Excising Drug Selection Cassette
Dickinson, D. J., Pani, A. M., Heppert, J. K., Higgins, C. D., & Goldstein, B. (2015) · Genetics · doi:10.1534/genetics.115.178335 · Cited by 586
CRISPR-Based Methods for Caenorhabditis elegans Genome Engineering
Dickinson, D. J., & Goldstein, B. (2016) · Genetics · doi:10.1534/genetics.115.182162 · Cited by 332
Precision genome editing using CRISPR-Cas9 and linear repair templates in C. elegans
Paix, A., Folkmann, A., & Seydoux, G. (2017) · Methods · doi:10.1016/j.ymeth.2017.03.023 · Cited by 256
Metadata
- Authors
- Ward, J. D.
- Venue
- Genetics
- Publisher
- Oxford University Press (OUP)
- Published
- 2014-12-09
- Volume
- 199
- Issue
- 2
- Pages
- 363-377
- ISSN
- 0016-6731 (ISSN-L)
- 1943-2631
- OpenAlex
- W2110819817
- Cited by
- 207
- References
- 6
- FWCI
- 6.45
- Type
- journal article
- Language
- en
- Open access
- green
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