Nigeria is a country located in Gulf of Guinea, in West Africa, facing the Atlantic Ocean. It is one of the most populous country in the world. There are more than 500 ethnic groups inhabiting the territory, representing a high diversity of languages and cultures. In this study, we collected 142 samples from the three largest ethnic groups in Nigeria: Hausa, Igbo and Yoruba. These samples were typed for the 27 Y-STR loci included in the Yfiler® Plus kit. A total of 140 different haplotypes were found, with two haplotypes shared by 2 individuals. The haplotype diversity was 0.9998 (±0.0009), slightly higher than that obtained with the 17 Y-STRs from the Yfiler kit (0.9992±0.0010). The Yfiler kit also showed an increased number of shared haplotypes, with 4 being shared by 2 individuals and one by 3. The average gene diversity over loci was lower in Nigeria than in the U.S. population groups reported in Yfiler® Plus kit User Guide. The DYS391, DYS390, DYS438, DYS392, DYS437 and DYS533 showed low diversity values (GD≤0.35). Comparisons between Nigeria and five populations in East Africa, the only African data available for the Yfiler® Plus markers, revealed significant differences among all populations (p-values ≪0.00005). The smallest FST was found with Kenya (FST=0.025), followed by Ethiopia (FST=0.105). As expected, distances with Djibouti, Eritrea and Somalia were higher that 20%, since our sample are from Bantu speakers, not represented in these population sample.