Fusing T7 exonuclease with the G-to-T base editor enhances the editing purity in rice

Qiurong Ren; Xiaojuan Gao; Tao Shi; Jiangmei Cao

doi:10.1016/j.jgg.2026.04.010

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Fusing T7 exonuclease with the G-to-T base editor enhances the editing purity in rice

doi: 10.1016/j.jgg.2026.04.010

a. Shanxi Key Laboratory of Nucleic Acid Biopesticides, School of Synthetic Biology, Shanxi University, Taiyuan, Shanxi 030006, China;
b. Institute of Biotechnology, Shanxi University, Taiyuan, Shanxi 030006, China

Funds:

This work was supported by the National Natural Science Foundation of China (award no. 32301248), the Fundamental Research Program of Shanxi Province (award no. 202303021212008), and the Wenying Young Scholars Talent Program of Shanxi University.

Received Date: 2025-12-14
Accepted Date: 2026-04-17
Rev Recd Date: 2026-04-14

Available Online: 2026-04-25

Abstract

Abstract

The G-to-T base editor (GTBE) is a genome editing tool that does not rely on deaminases. However, its application has been limited by low G-to-T editing efficiency and high insertion and deletion (InDel) byproduct rates. In this study, we develop a series of editors (GTBE1–GTBE7) using three strategies: modification of the Cas9 nickase (nCas9) variant, fusion with a transcriptional activation domain, and fusion with a 5′–3′ exonuclease targeting nicked DNA. Both protoplast and stable rice transformant analyses reveal that the GTBE7 editor, which incorporates T7 exonuclease, significantly improves G-to-T editing purity compared to the previously reported GTBE1 editor. In stable transformants, the InDel efficiency of GTBE7 is reduced by 14%–27.7% compared to GTBE1. To further broaden the editing range of GTBE7, we develop the GTBE7_nSpRY editor, enabling G-to-T base editing at non-NGG protospacer adjacent motif (PAM) sequences. Using GTBE7_nSpRY, we successfully create herbicide-resistant rice and identify the G628A mutation site, generating Imazethapyr-resistant rice plants through G-to-C byproduct edits. In summary, the GTBE7 and GTBE7_nSpRY editors developed in this study enhance G-to-T editing purity, reduce InDel byproduct efficiency, and offer potential for expanding the application of G-to-T base editing in plants.
- G-to-T base editor,
- T7 exonuclease,
- GTBE7,
- G-to-T editing purity,
- GTBE7_nSpRY

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References

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