This study aimed to determine gene pool structure in the tomato (Solanum lycopersicum) along with the relationships of populations in different regions. To do so, it leveraged 276 diverse cultivated accessions representing 35 countries, obtained via the USDA-GRIN tomato germplasm collection. Single nucleotide polymorphisms (SNPs) in these accessions were detected by sequencing, and genetic diversity, population structure, and phylogenetic relationships were estimated from the SNP alleles. The results of model-based ancestry analysis, phylogenetic tree construction, and principal component analysis supported these accessions as representing three well-differentiated genetic populations. Phylogenetic analyses traced the accessions back to their regions of origin, highlighting Central and South America as the primary center of domestication, with secondary centers in Europe and North America. This study provides a comprehensive analysis of the genetic variation in and relationships among cultivated tomato genotypes. The findings are of benefit to curators, researchers, and breeders in understanding, utilizing, conserving, and managing the collection, and in thereby contributing more efficiently to international tomato research.