Precise generation of bystander-free mouse models with ABE9-SpRY.

Ong, J. K.; Bhunia, S.; Hilbert, B.; Kirschner, V.; Dlugosz, S.; Zimmermann, F.; Freichel, M.; Cornean, A.

doi:10.1101/2025.10.07.680931

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bioRxiv · 10.1101/2025.10.07.680931

Precise generation of bystander-free mouse models with ABE9-SpRY.

Ong, J. K.·Bhunia, S.·Hilbert, B.·Kirschner, V.·Dlugosz, S.·Zimmermann, F.·Freichel, M.·Cornean, A.

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Abstract

Point mutations cause many genetic disorders, but modelling them in organisms is technically challenging. Creating mouse models that mimic these mutations is crucial for establishing a causal relationship between mutations and disease phenotype, thereby supporting the development of therapeutic strategies. Adenine base editors (ABEs) can correct single-nucleotide variants (SNVs) in disease modelling without double-stranded breaks (DSBs) or donor DNA, achieving higher product purity than traditional Cas9 methods. Earlier ABE techniques faced issues like limited targetability, bystander editing, and off-target effects. By combining two editor advancements, we introduced and tested ABE9-SpRY, an improved ABE variant fused with a PAM-flexible SpRY-Cas9 nickase. Our results show that ABE9-SpRY effectively generates three out of four targeted A-to-G mutations in mouse embryos, with significantly fewer off-target effects than ABE8e-SpRY, achieving desired editing efficiencies of up to 96% in individual adult founder mice.ABE9-SpRY also enhances product purity in mouse embryos and human induced pluripotent stem cells (hiPSCs) compared to ABE8e-SpRY. Our findings showcase ABE9-SpRYs precision and versatility, highlighting it as a powerful tool for accurate in vivo point mutation modelling.

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Ong, J. K., Bhunia, S., Hilbert, B., Kirschner, V., Dlugosz, S., Zimmermann, F., Freichel, M., Cornean, A.. 2025-10-07. Precise generation of bystander-free mouse models with ABE9-SpRY.. https://doi.org/10.1101/2025.10.07.680931

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