Genetius

The study generated a dataset of 420 sgRNAs targeting promoters, exons, and introns of 137 tomato genes in protoplasts, linking editing efficiency to chromatin accessibility, genomic context, and sequence features. Open chromatin sites showed higher editing rates, while transcriptional activity had little effect, and a subset of guides produced near‑complete editing with microhomology‑mediated deletions. Human‑trained prediction models performed poorly, highlighting the need for plant‑specific guide design tools.

Using a CRISPR/Cas9 multiplex system, the upstream regulatory region of the Ripening inhibitor (RIN) gene was edited to generate 264 T0 tomato lines with diverse promoter mutations and varying RIN transcription levels. The study uncovered a non‑linear relationship between promoter lesions and gene expression, setting the stage for future phenotypic and transcriptomic analyses of fruit quality.