A cytosine base editor toolkit with varying activity windows and target scopes for versatile gene manipulation in plants
Open accessAbstract
CRISPR/Cas-derived base editing tools empower efficient alteration of genomic cytosines or adenines associated with essential genetic traits in plants and animals. Diversified target sequences and customized editing products call for base editors with distinct features regarding the editing window and target scope. Here we developed a toolkit of plant base editors containing AID10, an engineered human AID cytosine deaminase. When fused to the N-terminus or C-terminus of the conventional Cas9 nickase (nSpCas9), AID10 exhibited a broad or narrow activity window at the protospacer adjacent motif (PAM)-distal and -proximal protospacer, respectively, while AID10 fused to both termini conferred an additive activity window. We further replaced nSpCas9 with orthogonal or PAM-relaxed Cas9 variants to widen target scopes. Moreover, we devised dual base editors with AID10 located adjacently or distally to the adenine deaminase ABE8e, leading to juxtaposed or spaced cytosine and adenine co-editing at the same target sequence in plant cells. Furthermore, we expanded the application of this toolkit in plants for tunable knockdown of protein-coding genes via creating upstream open reading frame and for loss-of-function analysis of non-coding genes, such as microRNA sponges. Collectively, this toolkit increases the functional diversity and versatility of base editors in basic and applied plant research.
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These single nucleotide polymorphisms (SNPs) can now be achieved through base or prime editing.
“These single nucleotide polymorphisms (SNPs) can now be achieved through base or prime editing (Lin et al., 2021;Xiong et al., 2022).”
From Genetic resources and precise gene editing for targeted improvement of barley abiotic stress tolerance (2023) · cites this paper as Xiong et al., 2022
Base editing can be repurposed as a gene loss of function tool by creating a stop codon or a gene knockdown by creating an upstream open reading frame (uORF).
“It can also be repurposed as a gene loss of function tool by creating a stop codon or a gene knockdown by creating an upstream open reading frame (uORF) (Billon et al., 2017;Kuscu et al., 2017;Xiong et al., 2022).”
From Genetic resources and precise gene editing for targeted improvement of barley abiotic stress tolerance (2023) · cites this paper as Xiong et al., 2022
Base-editing studies conducted in Arabidopsis and rice provide confidence in using this technique in barley to improve useful traits, for instance, inducing early or late flowering to escape stress windows such as those for frost or drought.
“Base-editing studies conducted in Arabidopsis and rice provide confidence in using this technique in barley to improve useful traits, for instance, inducing early or late flowering to escape stress windows such as those for frost or drought (Li et al., 2020;Xiong et al., 2022).”
From Genetic resources and precise gene editing for targeted improvement of barley abiotic stress tolerance (2023) · cites this paper as Xiong et al., 2022
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Systematic optimization of Cas12a base editors in wheat and maize using the ITER platform
Gaillochet, C., Fernández, A. P., Goossens, V., D’Halluin, K., Drozdzecki, A., Shafie, M., Duyse, J. V., Isterdael, G. V., Gonzalez, C., Vermeersch, M., et al. (2023) · Genome biology · doi:10.1186/s13059-022-02836-2 · Cited by 73 · see citing works
Progress in gene editing tools, implications and success in plants: a review
Bhuyan, S. J., Kumar, M., Devde, P. R., Rai, A. C., Mishra, A. K., Singh, P. K., & Siddique, K. H. M. (2023) · Frontiers in Genome Editing · doi:10.3389/fgeed.2023.1272678 · Cited by 59 · see citing works
Precise fine-turning of GhTFL1 by base editing tools defines ideal cotton plant architecture
Wang, G., Wang, F., Xu, Z., Wang, Y., Zhang, C., Zhou, Y., Hui, F., Yang, X., Nie, X., Zhang, X., et al. (2024) · Genome biology · doi:10.1186/s13059-024-03189-8 · Cited by 46 · see citing works
Liu, Q., Peng, X., Shen, M., Qian, Q., Xing, J., Li, C., & Gregory, R. I. (2022) · Nucleic Acids Research · doi:10.1093/nar/gkac1094 · Cited by 39 · see citing works
Self-citationLiang, Z., Yuan, L., Xiong, X., Hao, Y., Song, X., Zhu, T., Yu, Y., Fu, W., Lei, Y., Xu, J., et al. (2022) · The Plant Cell · doi:10.1093/plcell/koac217 · Cited by 36 · see citing works
Self-citationShelake, R. M., Pramanik, D., & Kim, J. (2023) · mBio · doi:10.1128/mbio.02296-22 · Cited by 33 · see citing works
Tian, J., Tang, Z., Niu, R., Zhou, Y., Yang, D., Chen, D., Luo, M., Mou, R., Yuan, M., & Xu, G. (2024) · Science China Life Sciences · doi:10.1007/s11427-024-2588-9 · Cited by 32 · see citing works
Engineering crop performance with upstream open reading frames
Mou, R., Niu, R., Yang, R., & Xu, G. (2024) · Trends in Plant Science · doi:10.1016/j.tplants.2024.10.005 · Cited by 32 · see citing works
Promoter editing for the genetic improvement of crops
Shi, L., Su, J., Cho, M., Song, H., Dong, O. X., Liang, Y., & Zhang, Z. (2023) · Journal of Experimental Botany · doi:10.1093/jxb/erad175 · Cited by 33 · see citing works
Split complementation of base editors to minimize off-target edits
Xiong, X., Liu, K., Li, Z., Xia, F., Ruan, X.-M., He, X., & Li, J. (2023) · Nature Plants · doi:10.1038/s41477-023-01540-8 · Cited by 32 · see citing works
Self-citationCapdeville, N., Schindele, P., & Puchta, H. (2022) · Current Opinion in Biotechnology · doi:10.1016/j.copbio.2022.102854 · Cited by 31 · see citing works
Plant genome editing: CRISPR, base editing, prime editing, and beyond
Xie, Y., Haq, S. I. U., Jiang, X., Zheng, D., Feng, N., Wang, W., He, J., & Qiu, Q. (2022) · Grassland Research · doi:10.1002/glr2.12034 · Cited by 27 · see citing works
Recent advances in therapeutic CRISPR-Cas9 genome editing: mechanisms and applications
Zhou, L., & Yao, S. (2023) · Molecular Biomedicine · doi:10.1186/s43556-023-00115-5 · Cited by 27 · see citing works
Efficient and heritable A-to-K base editing in rice and tomato
Li, X., Xie, J., Dong, C., Zheng, Z. S., Shen, R., Cao, X., Chen, X., Wang, M., Zhu, J., & Tian, Y. (2023) · Horticulture Research · doi:10.1093/hr/uhad250 · Cited by 25 · see citing works
Lokya, V., Singh, S., Chaudhary, R., Jangra, A., & Tiwari, S. (2025) · Plant Molecular Biology · doi:10.1007/s11103-025-01600-x · Cited by 20 · see citing works
Karunarathne, S., Walker, E., Sharma, D., Li, C., & Han, Y. (2023) · Journal of Zhejiang University-SCIENCE B · doi:10.1631/jzus.b2200552 · Cited by 18 · see citing works
“Base-editing studies conducted in Arabidopsis and rice provide confidence in using this technique in barley to improve useful traits, for instance, inducing early or late flowering to escape stress windows such as those for frost or drought (Li et al., 2020; Xiong et al., 2022).”
Self-citationShoot-root signal circuit: Phytoremediation of heavy metal contaminated soil
Bai, S., Han, X., & Feng, D. (2023) · Frontiers in Plant Science · doi:10.3389/fpls.2023.1139744 · Cited by 19 · see citing works
Genome editing for plant synthetic metabolic engineering and developmental regulation
Tan, J., Shen, M., Chai, N., Liu, Q., Liu, Y., & Zhu, Q. (2023) · Journal of Plant Physiology · doi:10.1016/j.jplph.2023.154141 · Cited by 16 · see citing works
A nuclease-dead Cas9-derived tool represses target gene expression
Wang, B., Liu, X., Li, Z., Zeng, K., Guo, J., Xin, T., Zhang, Z., Li, J., & Yang, X. Y. (2024) · PLANT PHYSIOLOGY · doi:10.1093/plphys/kiae149 · Cited by 15 · see citing works
Self-citationIn‐locus gene silencing in plants using genome editing
Shen, R., Yao, Q., Tan, X., Ren, W., Zhong, D., Zhang, X., Li, X., Dong, C., Cao, X., Tian, Y., et al. (2024) · New Phytologist · doi:10.1111/nph.19856 · Cited by 15 · see citing works
Berríos, K. N., Barka, A., Gill, J., Serrano, J. C., Bailer, P., Parker, J. B., Evitt, N. H., Gajula, K. S., Shi, J., & Kohli, R. M. (2024) · Nucleic Acids Research · doi:10.1093/nar/gkae024 · Cited by 12 · see citing works
CRISPR base editor-based targeted random mutagenesis (BE-TRM) toolbox for directed evolution
Shelake, R. M., Pramanik, D., & Kim, J. (2024) · BMB Reports · doi:10.5483/bmbrep.2023-0086 · Cited by 11 · see citing works
Zhong, Z., Hu, X., Zhang, R., Liu, X., Chen, W., Zhang, S., Sun, J., & Zhong, T. P. (2024) · Journal of genetics and genomics/Journal of Genetics and Genomics · doi:10.1016/j.jgg.2024.10.006 · Cited by 9 · see citing works
Cytosine base editors optimized for genome editing in potato protoplasts
Westberg, I. N., Carlsen, F. M., Johansen, I. E., & Petersen, B. L. (2023) · Frontiers in Genome Editing · doi:10.3389/fgeed.2023.1247702 · Cited by 10 · see citing works
Prasad, K. V. S. K., Cheema, A., Scanlon, W., Matthews, A., Sharifova, S., Huq, E., & Reddy, A. S. N. (2024) · PLANT PHYSIOLOGY · doi:10.1093/plphys/kiae396 · Cited by 8 · see citing works
Simoni, S., Fambrini, M., Pugliesi, C., & Rogo, U. (2025) · International Journal of Molecular Sciences · doi:10.3390/ijms26199294 · Cited by 7 · see citing works
Knockout of phosphatidate phosphohydrolase genes confers broad‐spectrum disease resistance in plants
Gong, Q., Sha, G., Han, X., Guo, Z., Yang, L., Chen, T., Yang, W., Tan, R., Liu, M., Xia, F., et al. (2024) · Plant Biotechnology Journal · doi:10.1111/pbi.14477 · Cited by 5 · see citing works
Wang, F., Bao, Y., Yao, X., Gong, B., Xiong, X., Wang, J., Li, Z., & Li, J. (2026) · Science China Life Sciences · doi:10.1007/s11427-025-3119-0 · Cited by 4 · see citing works
Self-citationSystematic optimization of Cas12a base editors in wheat and maize using the ITER platformPreprint
Gaillochet, C., Fernández, A. P., Goossens, V., D’Halluin, K., Drozdzecki, A., Shafie, M., Duyse, J. V., Isterdael, G. V., Gonzalez, C., Vermeersch, M., et al. (2022) · bioRxiv (Cold Spring Harbor Laboratory) · doi:10.1101/2022.05.11.491340 · Cited by 4 · see citing works
Chen, N., Gao, M., Bai, Y., Wang, M., Liu, M., Xiong, W., & Mo, B. (2026) · PLANT PHYSIOLOGY · doi:10.1093/plphys/kiag073 · Cited by 3 · see citing works
Efforts to Downsize Base Editors for Clinical Applications
Song, B. (2025) · International Journal of Molecular Sciences · doi:10.3390/ijms26052357 · Cited by 3 · see citing works
Versatile Applications of CRISPR ‐Based Programmable T‐ DNA Integration in Plants
Lin, X., Gong, B., Wang, F., Wan, J., Xiong, X., & Li, J. (2025) · Plant Biotechnology Journal · doi:10.1111/pbi.70353 · Cited by 3 · see citing works
Self-citationModular cytosine base editing promotes epigenomic and genomic modifications
Weischedel, J., Higgins, L., Rogers, S. L., Gramalla-Schmitz, A., Wyrzykowska, P., Borgoni, S., MacCarthy, T., & Chahwan, R. (2023) · Nucleic Acids Research · doi:10.1093/nar/gkad1118 · Cited by 1 · see citing works
Precision genetic manipulation: CRISPR toolkits reshape plant biology
Li, Z., Xiong, X., Cao, R., & Li, J. (2026) · PLANT PHYSIOLOGY · doi:10.1093/plphys/kiag696 · Cited by 1 · see citing works
Self-citationProgress in developing and implementing CRISPR technologies made by Sun Yat-sen University
Li, Z., Zhou, Y., Huang, J., & Li, J. (2024) · Scientia Sinica Vitae · doi:10.1360/ssv-2024-0145
Self-citationEpigenetics for Combating Plant Pathogens: Updated Methods and Current Achievements
Kouighat, M., Farhaoui, A., Assouguem, A., Taoussi, M., Belabess, Z., & Lahlali, R. (2025) · CABI climate change series. · doi:10.1079/9781800626102.0018
Gaillochet, C., Fernández, A. P., Goossens, V., D’Halluin, K., Drozdzecki, A., Shafie, M., Van Duyse, J., Isterdael, G. V., Gonzalez, C., Vermeersch, M., et al. (2023) · Figshare · doi:10.6084/m9.figshare.22611368
Gaillochet, C., Fernández, A. P., Goossens, V., D’Halluin, K., Drozdzecki, A., Shafie, M., Van Duyse, J., Isterdael, G. V., Gonzalez, C., Vermeersch, M., et al. (2023) · Figshare · doi:10.6084/m9.figshare.22611395
Engineering ADAR1p150- and dCasRx-based systems for programmable RNA base editing in plants
Ruan, X.-M., Zhang, Z.-Y., Xiong, X., Li, Y., Li, X., Li, J., & Wang, F. (2026) · Biochemical and Biophysical Research Communications · doi:10.1016/j.bbrc.2026.154603
Self-citationNano Delivery Systems for In Vivo CRISPR/Cas-Based Gene Therapy: From Vectors to Clinical Outcomes
Chen, R., Zhou, H., Wu, Q., & Gong, C. (2026) · ACS Nano · doi:10.1021/acsnano.6c12154
From base transitions to expanded editing tools: development and applications of plant base editors
Sui, N., Zhang, J., Xu, W., 庞美新, Zhang, R., Chai, N., Wen, J., Liu, T., Zheng, Z., Huang, J., et al. (2026) · Biotechnology Advances · doi:10.1016/j.biotechadv.2026.109074
- Authors
- Xiong, X., Li, Z., Liang, J., Liu, K., Li, C., & Li, J.-F.
- Affiliations
- Sun Yat-sen University (CN)
- MOE Key Laboratory of Gene Function and Regulation, State Key Laboratory of Biocontrol, Guangdong Provincial Key Laboratory of Plant Resources, School of Life Sciences, Sun Yat-sen University, Guangzhou 510275, China
- Corresponding author
- Li, Zhenxiang
- Venue
- Nucleic Acids Research
- Publisher
- Oxford University Press (OUP)
- Published
- 2022-03-14
- Volume
- 50
- Issue
- 6
- Pages
- 3565-3580
- ISSN
- 0305-1048 (ISSN-L)
- 1362-4962
- PMC
- PMC8989527
- OpenAlex
- W4214533861
- Cited by
- 41
- References
- 86
- Citations by year
- 2026: 6 · 2025: 5 · 2024: 11 · 2023: 14 · 2022: 5
- FWCI
- 2.85
- Type
- journal article
- Language
- en
- Open access
- gold
- doi.org
- Licence
- Locations
- Nucleic Acids Research (publishedVersion, cc-by-nc)
- PubMed Central (publishedVersion, cc-by-nc)
- PubMed (publishedVersion)
- Funders
- National Transgenic Science and Technology Major Program of China (2019ZX08010003-001-009)
- Foundation of Guangzhou Science and Technology Key Project (201904020041)
- National Natural Science Foundation of China (32125004)
- China Postdoctoral Science Foundation (2020TQ0387)
- Topics
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › CRISPR and Genetic Engineering
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › RNA regulation and disease
- Life Sciences › Agricultural and Biological Sciences › Plant Science › Plant Molecular Biology Research
- Keywords
- cytosine base editing, protospacer adjacent motif, ABE8e, plant genome editing, microRNA sponge
- MeSH terms
- Gene Editing, CRISPR-Associated Protein 9, Adenine, Animals, Cytosine, CRISPR-Cas Systems
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