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6. Genome-edited foods

Author

Tuncel, Aytug, Pan, Changtian, Sprink, Thorben, Wilhelm, Ralf, Barrangou, Rodolphe, Li, Li, Shih, Patrick M., Varshney, Rajeev K., Tripathi, Leena, Van Eck, Joyce, Mandadi, Kranthi, and Qi, Yiping

Published
2023
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7. Heritable base-editing in Arabidopsis using RNA viral vectors

Author

Liu, Degao, Xuan, Shuya, Prichard, Lynn E, Donahue, Lilee I, Pan, Changtian, Nagalakshmi, Ugrappa, Ellison, Evan E, Starker, Colby G, Dinesh-Kumar, Savithramma P, Qi, Yiping, and Voytas, Daniel F

Subjects
Arabidopsis, Arabidopsis Proteins, Mutation, RNA, RNA Editing, RNA, Plant, Biological Sciences, Agricultural and Veterinary Sciences, Plant Biology & Botany
Abstract

Heritable base-editing using a viral delivery system enables high-throughput functional analysis of genes in Arabidopsis.

Published
2022

21. PAM‐relaxed and temperature‐tolerant CRISPR‐Mb3Cas12a single transcript unit systems for efficient singular and multiplexed genome editing in rice, maize, and tomato.

Author

Liu, Shishi, He, Yao, Fan, Tingting, Zhu, Meirui, Qi, Caiyan, Ma, Yanqin, Yang, Mengqiao, Yang, Liang, Tang, Xu, Zhou, Jianping, Zhong, Zhaohui, An, Xueli, Qi, Yiping, and Zhang, Yong

Subjects
PLANT genomes, GENE expression, PROMOTERS (Genetics), ASPARTIC acid, GENETIC transcription, CRISPRS, GENOME editing
Abstract

Summary: Class 2 Type V‐A CRISPR‐Cas (Cas12a) nucleases are powerful genome editing tools, particularly effective in A/T‐rich genomic regions, complementing the widely used CRISPR‐Cas9 in plants. To enhance the utility of Cas12a, we investigate three Cas12a orthologs—Mb3Cas12a, PrCas12a, and HkCas12a—in plants. Protospacer adjacent motif (PAM) requirements, editing efficiencies, and editing profiles are compared in rice. Among these orthologs, Mb3Cas12a exhibits high editing efficiency at target sites with a simpler, relaxed TTV PAM which is less restrictive than the canonical TTTV PAM of LbCas12a and AsCas12a. To optimize Mb3Cas12a, we develop an efficient single transcription unit (STU) system by refining the linker between Mb3Cas12a and CRISPR RNA (crRNA), nuclear localization signal (NLS), and direct repeat (DR). This optimized system enables precise genome editing in rice, particularly for fine‐tuning target gene expression by editing promoter regions. Further, we introduced Arginine (R) substitutions at Aspartic acid (D) 172, Asparagine (N) 573, and Lysine (K) 579 of Mb3Cas12a, creating two temperature‐tolerant variants: Mb3Cas12a‐R (D172R) and Mb3Cas12a‐RRR (D172R/N573R/K579R). These variants demonstrate significantly improved editing efficiency at lower temperatures (22 °C and 28 °C) in rice cells, with Mb3Cas12a‐RRR showing the best performance. We extend this approach by developing efficient Mb3Cas12a‐RRR STU systems in maize and tomato, achieving biallelic mutants targeting single or multiple genes in T0 lines cultivated at 28 °C and 25 °C, respectively. This study significantly expands Cas12a's targeting capabilities in plant genome editing, providing valuable tools for future research and practical applications. [ABSTRACT FROM AUTHOR]

Published
2025
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22. An efficient CRISPR‐Cas12a‐mediated MicroRNA knockout strategy in plants.

Author

Zheng, Xuelian, Tang, Xu, Wu, Yuechao, Zheng, Xiaoqin, Zhou, Jianping, Han, Qinqin, Tang, Yalan, Fu, Xinxuan, Deng, Jiao, Wang, Yibo, Wang, Danning, Zhang, Shuting, Zhang, Tao, Qi, Yiping, and Zhang, Yong

Subjects
GENE expression, PLANT genes, GENOME editing, SEED development, PHENOTYPES
Abstract

Summary: In recent years, the CRISPR‐Cas9 nuclease has been used to knock out MicroRNA (miRNA) genes in plants, greatly promoting the study of miRNA function. However, due to its propensity for generating small insertions and deletions, Cas9 is not well‐suited for achieving a complete knockout of miRNA genes. By contrast, CRISPR‐Cas12a nuclease generates larger deletions, which could significantly disrupt the secondary structure of pre‐miRNA and prevent the production of mature miRNAs. Through the case study of OsMIR390 in rice, we confirmed that Cas12a is a more efficient tool than Cas9 in generating knockout mutants of a miRNA gene. To further demonstrate CRISPR‐Cas12a‐mediated knockout of miRNA genes in rice, we targeted nine OsMIRNA genes that have different spaciotemporal expression and have not been previously investigated via genetic knockout approaches. With CRISPR‐Cas12a, up to 100% genome editing efficiency was observed at these miRNA loci. The resulting larger deletions suggest Cas12a robustly generated null alleles of miRNA genes. Transcriptome profiling of the miRNA mutants, as well as phenotypic analysis of the rice grains revealed the function of these miRNAs in controlling gene expression and regulating grain quality and seed development. This study established CRISPR‐Cas12a as an efficient tool for genetic knockout of miRNA genes in plants. [ABSTRACT FROM AUTHOR]

Published
2025
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27. Foundational and Translational Research Opportunities to Improve Plant Health.

Author

Michelmore, Richard, Coaker, Gitta, Bart, Rebecca, Beattie, Gwyn, Bent, Andrew, Bruce, Toby, Cameron, Duncan, Dangl, Jeffery, Dinesh-Kumar, Savithramma, Edwards, Rob, Eves-van den Akker, Sebastian, Gassmann, Walter, Greenberg, Jean T, Hanley-Bowdoin, Linda, Harrison, Richard J, Harvey, Jagger, He, Ping, Huffaker, Alisa, Hulbert, Scot, Innes, Roger, Jones, Jonathan DG, Kaloshian, Isgouhi, Kamoun, Sophien, Katagiri, Fumiaki, Leach, Jan, Ma, Wenbo, McDowell, John, Medford, June, Meyers, Blake, Nelson, Rebecca, Oliver, Richard, Qi, Yiping, Saunders, Diane, Shaw, Michael, Smart, Christine, Subudhi, Prasanta, Torrance, Lesley, Tyler, Bret, Valent, Barbara, and Walsh, John

Subjects
Microbiology, Plant Biology, Biological Sciences, Genetics, Zero Hunger, Agriculture, Biotechnology, Climate Change, Crops, Agricultural, Food Supply, Humans, Plant Diseases, Translational Research, Biomedical, Plant Biology & Botany, Plant biology
Abstract

Reader Comments | Submit a Comment The white paper reports the deliberations of a workshop focused on biotic challenges to plant health held in Washington, D.C. in September 2016. Ensuring health of food plants is critical to maintaining the quality and productivity of crops and for sustenance of the rapidly growing human population. There is a close linkage between food security and societal stability; however, global food security is threatened by the vulnerability of our agricultural systems to numerous pests, pathogens, weeds, and environmental stresses. These threats are aggravated by climate change, the globalization of agriculture, and an over-reliance on nonsustainable inputs. New analytical and computational technologies are providing unprecedented resolution at a variety of molecular, cellular, organismal, and population scales for crop plants as well as pathogens, pests, beneficial microbes, and weeds. It is now possible to both characterize useful or deleterious variation as well as precisely manipulate it. Data-driven, informed decisions based on knowledge of the variation of biotic challenges and of natural and synthetic variation in crop plants will enable deployment of durable interventions throughout the world. These should be integral, dynamic components of agricultural strategies for sustainable agriculture.

Published
2017

37. Expanding the scope of plant genome engineering with Cas12a orthologs and highly multiplexable editing systems

Author

Zhang, Yingxiao, Ren, Qiurong, Tang, Xu, Liu, Shishi, Malzahn, Aimee A., Zhou, Jianping, Wang, Jiaheng, Yin, Desuo, Pan, Changtian, Yuan, Mingzhu, Huang, Lan, Yang, Han, Zhao, Yuxin, Fang, Qing, Zheng, Xuelian, Tian, Li, Cheng, Yanhao, Le, Ysa, McCoy, Bailey, Franklin, Lidiya, Selengut, Jeremy D., Mount, Stephen M., Que, Qiudeng, Zhang, Yong, and Qi, Yiping

Published
2021
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48. Broad range plastid genome editing with monomeric TALE‐linked cytosine and dual base editors.

Author

Wang, Xiaoyu, Fang, Tyson, Lu, Jason, Tripathi, Leena, and Qi, Yiping

Subjects
RNA polymerase II, GENOME editing, CYTIDINE deaminase, PLANT mitochondria, TRANSGENIC rice
Abstract

This article discusses the development of new monomeric TALE-linked base editors for editing the plastid genome in rice. The researchers explored different cytidine deaminases to generate these base editors, including a human APOBEC3A variant and an improved cytidine deaminase based on TadA. They evaluated the editing efficiency of these base editors in rice calli and transgenic rice plants, finding that the monomeric TALE-linked base editors induced high-efficiency C-to-T conversions. The researchers also examined the off-target activity of these base editors and discussed the advantages they offer over other base editors. Overall, this study demonstrates the potential of these monomeric TALE-linked base editors for efficient plastid genome editing in rice. [Extracted from the article]

Published
2024
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