The purpose of this study was to identify paternity relations and increase the likelihood success in DNA identification of biological samples found in mass disasters and criminal events by using the 6-dye GlobalFilerTM amplification kit with 21 autosomal STR loci in our laboratory for validation purpose and routine use. In this regard, swab samples from different regions of Turkey (n=350) were collected and studied. The validation of the GlobalFilerTM Amplification Kit was carried out in accordance with the guidelines published by the Scientific Working Group on DNA Analysis Methods (SWGDAM). Our study was conducted to show the efficacy and performance of the kit for sensitivity, specificity, limit of detection, dynamic range, limit of quantification, stochastic threshold study, reproducibility and repeatability, mixture and contamination study parameters. Validation parameters and population genetics data were calculated by using Arlequin v 22.214.171.124 version. According to our results, SE33 (PM=0.014) locus showed the greatest power of discrimination and TPOX (PM=0.132) has the least power of discrimination power in Turkish population. Combined power of exclusion and combined power of discrimination were calculated as 99.99999963% and 99.99999999%, respectively. In addition, allelic frequencies from our study were compared to previously published population data of Iraq, Saud Arabia, China (Han), USA (Cauc), Iran, Afghan, Azerbaijan, Romania, and South Africa. The study revealed that there was a significant difference between Turkish and South Africa (0.1198) population and no difference was observed for Azerbaijan (0.0097) population.