Genome- and transcriptome-wide off-target analyses of a high-efficiency adenine base editor in tomato

Simon Sretenovic; Yumi Green; Yuechao Wu; Yanhao Cheng; Tao Zhang; Joyce Van Eck; Yiping Qi

doi:10.1093/plphys/kiad347

journal article Jun 14, 2023

Genome- and transcriptome-wide off-target analyses of a high-efficiency adenine base editor in tomato

Simon Sretenovic

Yumi Green Yuechao Wu

Yanhao Cheng Tao Zhang

Joyce Van Eck

Yiping Qi

Plant Physiology Vol. 193 No. 1 pp. 291-303 · Oxford University Press (OUP)

View at Publisher Save 10.1093/plphys/kiad347

Abstract

Abstract
Adenine base editors (ABEs) are valuable, precise genome editing tools in plants. In recent years, the highly promising ADENINE BASE EDITOR8e (ABE8e) was reported for efficient A-to-G editing. However, compared to monocots, comprehensive off-target analyses for ABE8e are lacking in dicots. To determine the occurrence of off-target effects in tomato (Solanum lycopersicum), we assessed ABE8e and a high-fidelity version, ABE8e-HF, at 2 independent target sites in protoplasts, as well as stable T0 lines. Since ABE8e demonstrated higher on-target efficiency than ABE8e-HF in tomato protoplasts, we focused on ABE8e for off-target analyses in T0 lines. We conducted whole-genome sequencing (WGS) of wild-type (WT) tomato plants, green fluorescent protein (GFP)–expressing T0 lines, ABE8e-no-gRNA control T0 lines, and edited T0 lines. No guide RNA (gRNA)–dependent off-target edits were detected. Our data showed an average of approximately 1,200 to 1,500 single-nucleotide variations (SNVs) in either GFP control plants or base-edited plants. Also, no specific enrichment of A-to-G mutations were found in base-edited plants. We also conducted RNA sequencing (RNA-seq) of the same 6 base-edited and 3 GFP control T0 plants. On average, approximately 150 RNA–level SNVs were discovered per plant for either base-edited or GFP controls. Furthermore, we did not find enrichment of a TA motif on mutated adenine in the genomes and transcriptomes in base-edited tomato plants, as opposed to the recent discovery in rice (Oryza sativa). Hence, we could not find evidence for genome- and transcriptome-wide off-target effects by ABE8e in tomato.

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Details

Published: Jun 14, 2023
Vol/Issue: 193(1)
Pages: 291-303
License: View

Authors

S

Simon Sretenovic

Department of Plant Science and Landscape Architecture, University of Maryland , College Park, MD 20742 , USA

Y

Yumi Green

The Boyce Thompson Institute , Ithaca, NY 14853 , USA

Y

Yuechao Wu

Jiangsu Key Laboratory of Crop Genomics and Molecular Breeding/Key Laboratory of Plant Functional Genomics of the Ministry of Education, College of Agriculture, Yangzhou University , Yangzhou 225009 , China; Jiangsu Co-Innovation Center for Modern Production Technology of Grain Crops, Yangzhou University , Yangzhou 225009 , China

Y

Yanhao Cheng

Department of Plant Science and Landscape Architecture, University of Maryland , College Park, MD 20742 , USA

T

Tao Zhang

Jiangsu Key Laboratory of Crop Genomics and Molecular Breeding/Key Laboratory of Plant Functional Genomics of the Ministry of Education, College of Agriculture, Yangzhou University , Yangzhou 225009 , China; Jiangsu Co-Innovation Center for Modern Production Technology of Grain Crops, Yangzhou University , Yangzhou 225009 , China

J

Joyce Van Eck

The Boyce Thompson Institute , Ithaca, NY 14853 , USA; Plant Breeding and Genetics Section, Cornell University , Ithaca, NY 14853 , USA

Y

Yiping Qi

Department of Plant Science and Landscape Architecture, University of Maryland , College Park, MD 20742 , USA; Institute for Bioscience and Biotechnology Research, University of Maryland , Rockville, MD 20850 , USA

Funding

U.S. Department of Agriculture Biotechnology Risk Assessment Grant Program Award: 2018-33522-28789

National Science Foundation Plant Genome Research Program Award: IOS-2029889

Foundation for Food and Agriculture Research Fellow

Cite This Article

Simon Sretenovic, Yumi Green, Yuechao Wu, et al. (2023). Genome- and transcriptome-wide off-target analyses of a high-efficiency adenine base editor in tomato. Plant Physiology, 193(1), 291-303. https://doi.org/10.1093/plphys/kiad347

Genome- and transcriptome-wide off-target analyses of a high-efficiency adenine base editor in tomato

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