CRISPR-mediated gap-based replacement enables site-specific DNA lesion insertion into long DNA and nucleosomes.

CGRID enables efficient, site-specific insertion of diverse DNA lesions into long DNA and nucleosomes, supporting high-throughput single-molecule repair analysis.

• Why it matters: Understanding DNA repair mechanisms requires precise lesion placement, but current methods lack flexibility, efficiency, or compatibility with chromatinized DNA, limiting mechanistic insights.
• What they did: The study developed CGRID, a programmable approach using paired Cas9 nickases to generate gaps and replace them with lesion-containing oligonucleotides, applied to λ-DNA and nucleosomes.
• The result: CGRID allows efficient installation of single lesions, double-strand breaks, and damage repositioning within nucleosomes, revealing that nucleosome context and lesion orientation influence repair enzyme activity.