Cultivated tomato (Solanum lycopersicum L.) is an economically important vegetable and has a narrow genetic base due to intensive human selection through domestication and breeding. The low level of genetic variation between cultivated tomatoes has made it difficult to develop molecular markers for elite breeding lines. Recently, genome-wide 145,695 InDels were identified from in silico analysis of two tomato genome sequences, Heinz 1706 (S. lycoperiscum) and LA1589 (S. pimpnellifolium). Of these, 2,272 InDels were validated and 717 InDels showed polymorphism in cultivated tomatoes. In the present study, we selected 48 out of 717 InDels based on PIC value (> 0.3) and size (> 10 bp) to develop a DNA database for commercial tomato cultivars. We also used an additional set of 28 InDels that have been previously reported. These markers were distributed across 11 chromosomes with an average of 6.6 markers. A total of 48 F1 hybrid cultivars were collected from 20 seed companies and a subset of eight cultivars were used to test polymorphism of the InDel markers. The 37 InDel markers were polymorphic in these cultivars and were used to genotype additional 40 cultivars. Genetic distances and relationships between cultivars were assessed using the InDel genotypes of 48 cultivars. This analysis revealed that the InDel markers detected genetic variations to identify 46 cultivars. Our results demonstrate that the InDel markers will be a useful resource to construct a DNA database for tomato cultivars and to protect tomato breeder’s rights via variety identification.