The aim of this study was to evaluate phenotypic traits and genetic diversity of the 13 tomato (Solanum lycopersicum L.) varieties and 6 hybrids developed at the Institute of Horticulture Lithuanian Research Centre for Agriculture and Forestry (LRCAF IH). For the molecular characterisation, seven previously published microsatellite markers (SSR) were used. A24 and 26 alleles were detected in tomato varieties and hybrids, respectively. Based on the polymorphism information content (PIC) value, the most informative SSR primers for varieties were TMS52, TGS0007, LEMDDNa and Tom236-237, and the most informative SSR primers for hybrids were SSR248 and TMS52. In UPGMA cluster analysis, tomato varieties are grouped in some cases due to genetic relationships, as the same cluster cultivars ‘Viltis’ (the parent of cv. ‘Laukiai’) and ‘Aušriai’ (the progeny of cv. ‘Jurgiai’) are present. The grouping of all hybrids in the dendrogram is related to the parental forms, and it shows the usefulness of molecular markers for tomato breeding, as they can be used to trace the origin of hybrids and, eventually, varieties accurately. The knowledge about the genetic background of Lithuanian tomato cultivars will help plan targeted crosses in tomato breeding programs.