What controls establish that a gene-editing experiment worked as intended?

Why

Begin with the intended allele: knockout, precise substitution or insertion. Sequence the target locus using a method that resolves mixed genotypes, and inspect both alleles or subclones when interpretation requires it. A non-targeting guide and unedited parental cells reveal process effects.

Independent guides, a rescue construct or orthogonal perturbation strengthen causal attribution. For a knockout, check protein loss and a target-specific functional readout, because an indel may preserve an open reading frame or an alternative isoform may remain. For a precise edit, verify the donor-derived substitution and unintended nearby insertions or deletions.

Off-target analysis should be proportionate to the use case, without claiming whole-genome certainty from one assay. For instance, a shifted PCR band does not establish that both copies of a receptor gene are inactive. Document clone selection and population heterogeneity, which can introduce additional confounding.