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Method: High-throughput and genome-scale targeted mutagenesis using CRISPR in a nonmodel multicellular organism, Bombyx mori
- Sanyuan Ma,
- Tong Zhang,
- Ruolin Wang,
- Pan Wang,
- Yue Liu,
- Jiasong Chang,
- Aoming Wang,
- Xinhui Lan,
- Le Sun,
- Hao Sun,
- Run Shi,
- Wei Lu,
- Dan Liu,
- Na Zhang,
- Wenbo Hu,
- Xiaogang Wang,
- Weiqing Xing,
- Ling Jia,
- and Qingyou Xia
Genome Res. January 2024 34: 134-144; Published in Advance January 8, 2024, doi:10.1101/gr.278297.123
...to date. Here, we report the construction of a high-throughput CRISPR-Cas9 mutant library for B. mori, a useful resource for silkworm research and breeding. We show its application for gene function interrogation, as well as its potential use for genetic improvement of economic traits. In silkworm...
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Method: Structure-optimized sgRNA selection with PlatinumCRISPr for efficient Cas9 generation of knockouts
- Irmgard U. Haussmann,
- Thomas C. Dix,
- David W.J. McQuarrie,
- Veronica Dezi,
- Abdullah I. Hans,
- Roland Arnold,
- and Matthias Soller
Genome Res. December 2024 34: 2279-2292; Published in Advance December 3, 2024, doi:10.1101/gr.279479.124
...mismatch to the target gene in Drosophila (Ren et al. 2014), off-target scission has been reported in tissue culture cells (Fu et al. 2013; Hsu et al. 2013). To compare off-target effects with sgRNA cleavage efficiencies predicted by either PlatinumCRISPr, DeepSpCas9, or Azimuth (standard scoring...
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Review: A systematic guide for identifying transcription factors that directly regulate the expression of a gene of interest
- Andrew D. Bates,
- Dawid Grzela,
- Maciej Studzian,
- Louise Brennan,
- Moli Williams,
- Conor Fawcett,
- Beth Hammond,
- Manreen Grewal,
- Marcin Ratajewski,
- Lukasz Pulaski,
- and Urszula L. McClurg
Genome Res. March 2026 36: 433-459; Published in Advance February 17, 2026, doi:10.1101/gr.281154.125
...in incomplete knockdown and can suffer from off-target effects, which complicate interpretation of transcriptional changes.Genome editing approaches, such as CRISPR–Cas9-mediated knockout (Nakamura et al. 2021), enable permanent removal of a TF. Knockouts are powerful for defining essential regulatory roles...
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Resource: Systematic genome editing of the genes on zebrafish Chromosome 1 by CRISPR/Cas9
- Yonghua Sun,
- Bo Zhang,
- Lingfei Luo,
- De-Li Shi,
- Han Wang,
- Zongbin Cui,
- Honghui Huang,
- Ying Cao,
- Xiaodong Shu,
- Wenqing Zhang,
- Jianfeng Zhou,
- Yun Li,
- Jiulin Du,
- Qingshun Zhao,
- Jun Chen,
- Hanbing Zhong,
- Tao P. Zhong,
- Li Li,
- Jing-Wei Xiong,
- Jinrong Peng,
- Wuhan Xiao,
- Jian Zhang,
- Jihua Yao,
- Zhan Yin,
- Xianming Mo,
- Gang Peng,
- Jun Zhu,
- Yan Chen,
- Yong Zhou,
- Dong Liu,
- Weijun Pan,
- Yiyue Zhang,
- Hua Ruan,
- Feng Liu,
- Zuoyan Zhu,
- Anming Meng,
- and The ZAKOC Consortium
Genome Res. January 2020 30: 118-126; Published in Advance December 12, 2019, doi:10.1101/gr.248559.119
.... The flexibility of the CRISPR/Cas9 system and data from the completed zebrafish sequencing project make it possible to systematically knock out every single gene in zebrafish (Howe et al. 2013; Hwang et al. 2013; Varshney et al. 2015). However, properties of gene targeting via the CRISPR/Cas9...
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Method: Characterization of meiotic recombination intermediates through gene knockouts in founder hybrid mice
- Benjamin Davies,
- Gang Zhang,
- Daniela Moralli,
- Samy Alghadban,
- Daniel Biggs,
- Chris Preece,
- Peter Donnelly,
- and Anjali Gupta Hinch
Genome Res. November 2023 33: 2018-2027; Published in Advance November 17, 2023, doi:10.1101/gr.278024.123
...and electroporation of the F1 zygotes with CRISPR-Cas9 ribonucleoprotein targeting the Dmc1 gene. Offspring are putative biallelic knockout founders. DMC1 immunohistochemical analysis allows founders with mosaic or remnant DMC1 to be excluded from the analysis (represented as a hybrid founder mouse in parenthesis...
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Method: Highly efficient CRISPR/Cas9-mediated knock-in in zebrafish by homology-independent DNA repair
- Thomas O. Auer,
- Karine Duroure,
- Anne De Cian,
- Jean-Paul Concordet,
- and Filippo Del Bene
Genome Res. January 2014 24: 142-153; Published in Advance October 31, 2013, doi:10.1101/gr.161638.113
...the widespread application of gene targeting by HR because screening a large number of animals may be required to isolate founders carrying the mutation of interest. Here we report highly efficient CRISPR/Cas9-mediated knock-in of >5.7-kb-long DNA cassettes into the zebrafish based on homology-independent DSB...
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Method: Megabase-scale deletion using CRISPR/Cas9 to generate a fully haploid human cell line
- Patrick Essletzbichler,
- Tomasz Konopka,
- Federica Santoro,
- Doris Chen,
- Bianca V. Gapp,
- Robert Kralovics,
- Thijn R. Brummelkamp,
- Sebastian M.B. Nijman,
- and Tilmann Bürckstümmer
Genome Res. December 2014 24: 2059-2065; Published in Advance November 4, 2014, doi:10.1101/gr.177220.114
...of off-target editing. The use of the CRISPR/Cas9 system for deletion of specific genomic regions of interest paves the way for the functional characterization of promoters, enhancers, and other regulatory regions in the human . Moreover, the inactivation of entire gene clusters by deletion will enable...
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Method: Cre-dependent Cas9-expressing pigs enable efficient in vivo genome editing
- Kepin Wang,
- Qin Jin,
- Degong Ruan,
- Yi Yang,
- Qishuai Liu,
- Han Wu,
- Zhiwei Zhou,
- Zhen Ouyang,
- Zhaoming Liu,
- Yu Zhao,
- Bentian Zhao,
- Quanjun Zhang,
- Jiangyun Peng,
- Chengdan Lai,
- Nana Fan,
- Yanhui Liang,
- Ting Lan,
- Nan Li,
- Xiaoshan Wang,
- Xinlu Wang,
- Yong Fan,
- Pieter A. Doevendans,
- Joost P.G. Sluijter,
- Pentao Liu,
- Xiaoping Li,
- and Liangxue Lai
Genome Res. December 2017 27: 2061-2071; Published in Advance November 16, 2017, doi:10.1101/gr.222521.117
...representation of porcine EML4–ALK rearrangements induced by CRISPR-Cas9. EML4-sgRNA and ALK-sgRNA (red) were designed to target the mutation sites of the porcine EML4 gene intron 14 and porcine ALK gene intron 13. PCR primers are indicated (primers A, B, C, and D). (B) PCRs were performed to analyze ALK–EML4...
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Research: Genome wide analysis of 3′ UTR sequence elements and proteins regulating mRNA stability during maternal-to-zygotic transition in zebrafish
- Charles E. Vejnar,
- Mario Abdel Messih,
- Carter M. Takacs,
- Valeria Yartseva,
- Panos Oikonomou,
- Romain Christiano,
- Marlon Stoeckius,
- Stephanie Lau,
- Miler T. Lee,
- Jean-Denis Beaudoin,
- Damir Musaev,
- Hiba Darwich-Codore,
- Tobias C. Walther,
- Saeed Tavazoie,
- Daniel Cifuentes,
- and Antonio J. Giraldez
Genome Res. July 2019 29: 1100-1114; Published in Advance June 21, 2019, doi:10.1101/gr.245159.118
..., we used CRISPR-Cas9 editing to mutate a potential destabilizing sequence element identified in the trip10a gene. We observed that a 223-nt deletion in the endogenous gene, targeting the region regulated in RESA, caused stabilization of the mutant mRNA compared with the wild type, without any apparent...
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Method: Highly efficient gene knockout in mice and zebrafish with RNA-guided endonucleases
- Young Hoon Sung,
- Jong Min Kim,
- Hyun-Taek Kim,
- Jaehoon Lee,
- Jisun Jeon,
- Young Jin,
- Jung-Hwa Choi,
- Young Ho Ban,
- Sang-Jun Ha,
- Cheol-Hee Kim,
- Han-Woong Lee,
- and Jin-Soo Kim
Genome Res. January 2014 24: 125-131; Published in Advance November 19, 2013, doi:10.1101/gr.163394.113
...-guided endonucleases (RGENs), derived from the prokaryotic Type II CRISPR-Cas system, enable targeted modification in cells and organisms. Here we describe the establishment of gene-knockout mice and zebrafish by the injection of RGENs as Cas9 protein:guide RNA complexes or Cas9 mRNA plus guide RNA into one...

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