globalchange  > 气候变化与战略
DOI: 10.1073/pnas.2013338118
论文题名:
Efficient CRISPR-mediated base editing in agrobacterium spp
作者: Rodrigues S.D.; Karimi M.; Impens L.; van Lerberge E.; Coussens G.; Aesaert S.; Rombaut D.; Holtappels D.; Ibrahim H.M.M.; van Montagu M.; Wagemans J.; Jacobs T.B.; de Coninck B.; Pauwels L.
刊名: Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424
出版年: 2021
卷: 118, 期:2
语种: 英语
英文关键词: Agrobacterium ; Base editing ; CRISPR ; Hairy root disease ; Plant transformation
英文摘要: Agrobacterium spp. are important plant pathogens that are the causative agents of crown gall or hairy root disease. Their unique infection strategy depends on the delivery of part of their DNA to plant cells. Thanks to this capacity, these phytopathogens became a powerful and indispensable tool for plant genetic engineering and agricultural biotechnology. Although Agrobacterium spp. are standard tools for plant molecular biologists, current laboratory strains have remained unchanged for decades and functional gene analysis of Agrobacterium has been hampered by time-consuming mutation strategies. Here, we developed clustered regularly interspaced short palindromic repeats (CRISPR)-mediated base editing to enable the efficient introduction of targeted point mutations into the genomes of both Agrobacterium tumefaciens and Agrobacterium rhizogenes. As an example, we generated EHA105 strains with loss-of-function mutations in recA, which were fully functional for maize (Zea mays) transformation and confirmed the importance of RolB and RolC for hairy root development by A. rhizogenes K599. Our method is highly effective in 9 of 10 colonies after transformation, with edits in at least 80% of the cells. The genomes of EHA105 and K599 were resequenced, and genome-wide off-target analysis was applied to investigate the edited strains after curing of the base editor plasmid. The off-targets present were characteristic of Cas9-independent off-targeting and point to TC motifs as activity hotspots of the cytidine deaminase used. We anticipate that CRISPR-mediated base editing is the start of “engineering the engineer,” leading to improved Agrobacterium strains for more efficient plant transformation and gene editing. © 2021 National Academy of Sciences. All rights reserved.
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资源类型: 期刊论文
标识符: http://119.78.100.158/handle/2HF3EXSE/163348
Appears in Collections:气候变化与战略

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作者单位: Rodrigues, S.D., Division of Crop Biotechnics, Department of Biosystems, KU Leuven, Leuven, 3001, Belgium; Karimi, M., Department of Plant Biotechnology and Bioinformatics, Ghent University, Ghent, 9052, Belgium, Center for Plant Systems Biology, VIB, Ghent, 9052, Belgium; Impens, L., Department of Plant Biotechnology and Bioinformatics, Ghent University, Ghent, 9052, Belgium, Center for Plant Systems Biology, VIB, Ghent, 9052, Belgium; van Lerberge, E., Department of Plant Biotechnology and Bioinformatics, Ghent University, Ghent, 9052, Belgium, Center for Plant Systems Biology, VIB, Ghent, 9052, Belgium; Coussens, G., Department of Plant Biotechnology and Bioinformatics, Ghent University, Ghent, 9052, Belgium, Center for Plant Systems Biology, VIB, Ghent, 9052, Belgium; Aesaert, S., Department of Plant Biotechnology and Bioinformatics, Ghent University, Ghent, 9052, Belgium, Center for Plant Systems Biology, VIB, Ghent, 9052, Belgium; Rombaut, D., Department of Plant Biotechnology and Bioinformatics, Ghent University, Ghent, 9052, Belgium, Center for Plant Systems Biology, VIB, Ghent, 9052, Belgium; Holtappels, D., Laboratory of Gene Technology, Department of Biosystems, Division Animal and Human Health Engineering, KU Leuven, Leuven, 3001, Belgium; Ibrahim, H.M.M., Division of Crop Biotechnics, Department of Biosystems, KU Leuven, Leuven, 3001, Belgium; van Montagu, M., Department of Plant Biotechnology and Bioinformatics, Ghent University, Ghent, 9052, Belgium, Center for Plant Systems Biology, VIB, Ghent, 9052, Belgium, International Plant Biotechnology Outreach, VIB, Ghent, 9052, Belgium; Wagemans, J., Laboratory of Gene Technology, Department of Biosystems, Division Animal and Human Health Engineering, KU Leuven, Leuven, 3001, Belgium; Jacobs, T.B., Department of Plant Biotechnology and Bioinformatics, Ghent University, Ghent, 9052, Belgium, Center for Plant Systems Biology, VIB, Ghent, 9052, Belgium; de Coninck, B., Division of Crop Biotechnics, Department of Biosystems, KU Leuven, Leuven, 3001, Belgium; Pauwels, L., Department of Plant Biotechnology and Bioinformatics, Ghent University, Ghent, 9052, Belgium, Center for Plant Systems Biology, VIB, Ghent, 9052, Belgium

Recommended Citation:
Rodrigues S.D.,Karimi M.,Impens L.,et al. Efficient CRISPR-mediated base editing in agrobacterium spp[J]. Proceedings of the National Academy of Sciences of the United States of America,2021-01-01,118(2)
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