Synthetic gene drives based on CRISPR/Cas9 have the potential to control, alter or suppress populations of crop pests and disease vectors, but it is unclear how they will function in wild populations. Using genetic data from four populations of the flour beetle Tribolium castaneum, we show that most populations harbor genetic variants in Cas9 target sites, some of which would render them immune to drive (ITD). We show that even a rare ITD allele can reduce or eliminate the efficacy of CRISPR/Cas9-based synthetic gene. This effect is equivalent to and accentuated by mild inbreeding, which is a characteristic of many disease-vectoring arthropods. We conclude that designing such a drive will require characterization of genetic variability and the mating system within and among targeted populations.