CRISPR Gene Editing Simulator

CRISPR-Cas9 Gene Editing

gRNA specificity 0.85

Cas9 scan speed 1.0×

On-target cuts

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Off-target cuts

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Editing efficiency

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Repair pathway

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How it works

Cas9 is guided along the DNA by a short guide RNA (gRNA) whose sequence is complementary to the intended target. It only stops to check a site when a PAM motif (NGG) is present immediately downstream — without a PAM, Cas9 never engages even a perfect-match sequence. At a PAM site, the gRNA tries to base-pair with the local DNA: the true target matches fully, while a handful of "off-target" decoy sites nearby share partial homology. Lower specificity raises the chance the enzyme accepts those partial matches and cuts anyway, producing unwanted edits.

P(cut | site) = PAM(site) · [ site=target  OR  rand() < (1−specificity)·homology(site) ]
NHEJ: fast, no template → random indel → gene knock-out
HDR:  slow, needs donor template → precise edit → gene knock-in (lower efficiency)

gRNA specificity — how strictly the guide RNA rejects partial-match sequences; higher values suppress off-target cuts without changing on-target efficiency.
Repair pathway — after a double-strand break, the cell's repair machinery either patches it via error-prone NHEJ (fast, disrupts the gene — a knock-out) or copies a donor template via HDR (slower, only active in dividing cells, but precise enough to insert new sequence — a knock-in).
Cas9 scan speed — how quickly the Cas9-gRNA complex slides along the DNA searching for the next PAM candidate.
PAM sites — toggle the small flags marking every NGG motif Cas9 is willing to inspect, whether or not a cut ultimately happens there.

Real-world relevance: this specificity/repair-pathway trade-off is exactly what CRISPR therapeutics teams tune — base editing and prime editing were both developed to cut off-target risk further while keeping the precision HDR-style edits need.

Cas9 + gRNA Target site Off-target cut PAM site

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