Valorisation of tuber protein is relevant for the potato starch industry to create added-value and reduce impact on the environment. Hence, protein content has emerged as a key quality trait for innovative potato breeders. In this study, we estimated trait heritability, explored the relationship between protein content and tuber under-water weight (UWW), inferred haplotypes underlying quantitative trait loci (QTLs) and pinpointed candidate genes. We used a panel of varieties (N = 277) that was genotyped using the SolSTW 20 K Infinium single-nucleotide polymorphism (SNP) marker array. Protein content data were collected from multiple environments and years. Our genome-wide association study (GWAS) identified QTLs on chromosomes 3, 5, 7 and 12. Alleles of StCDF1 (maturity) were associated with QTLs found on chromosome 5. The QTLs on chromosomes 7 and 12 are presented here for the first time, whereas those on chromosomes 3 and 5 co-localized with loci reported in earlier studies. The candidate genes underlying the QTLs proposed here are relevant for functional studies. This study provides resources for genomics-enabled breeding for protein content in potato.