In CRISPR-Cas9 genome editing, how does the specificity of the Cas9 protein influence the success of gene editing?

The Cas9 protein must recognize a very specific DNA sequence called the protospacer adjacent motif (PAM) next to the target site; if it binds loosely, it can cut many off‑target places, causing unwanted mutations. When Cas9’s specificity is high, it cuts only the intended gene, leading to precise edits and fewer side effects. The success of gene editing therefore depends on how tightly Cas9 distinguishes the target sequence from similar ones, which is controlled by the guide RNA and the protein’s binding domain. For example, in a plant gene, a well‑designed guide RNA and Cas9 with high fidelity will change only the desired leaf‑color gene, while a less specific Cas9 might also alter a nearby growth‑rate gene, producing unpredictable traits. Thus, the more selective Cas9 is, the more reliable and safe the genome editing outcome becomes.