This dataset contains raw bioassay and molecular data from an insecticide resistance study of Anopheles gambiae sensu stricto populations in Ghana. WHO standard susceptibility tests were conducted using multiple insecticides, and 24-hour mortality was recorded to determine phenotypic resistance. Surviving mosquitoes were further analysed using PCR to detect kdr mutations in the voltage-gated sodium channel gene, with genotype frequencies recorded. The dataset supports a manuscript submitted to Malaria Journal and provides information on resistance status and underlying genetic mechanisms in malaria vectors from the study area.