Abstract The STRs are a widely used tool in forensic genetics and biological anthropology. This study established allele frequencies and some parameters of forensic interest for the autosomal markers included in the PowerPlex ® 16 BIO System kit (Promega). The sample population is from 151 unrelated afro-descendant individuals of San Basilio de Palenque (SBP), Colombia, a population that was founded for free Africans in the colony time and is considered the first free town in America. This population lacks published databases for these markers. DNA was isolated by salting-out. The products were amplified with the PowerPlex ® 16 BIO System kit (Promega) and separated by electrophoresis on 6% polyacrylamide gels and visualized with a FMBIO IIe Genetic Analyzer (HITACHI). Allelic diversity and Hardy-Weinberg equilibrium were performed using the ARLEQUIN software. Bonferroni correction assumes 0.05 significance level used for 15 tests yields an actual significance of 0.0033. The forensic parameters were calculated with PowerStats software (Promega CO). All markers analyzed showed heterozygosity from 66, 89% (TH01) until 87, 42% (Penta E and Penta D). All systems were in Hardy-Weinberg equilibrium after Bonferroni correction. The probabilities of paternity (W), exclusion (PE) and discrimination (PD) accumulated for loci analyzed were 0.99999899, 0.999999 and >0.99999999, respectively. The parameters of forensic interest had values suitable for routine use in forensic genetics.