The aim of this study was to evaluate the differences on motor competence between 5-year-old boys and girls and to investigate the existence of Relative Age Effect (RAE) on their motor competence. A total of 232 preschool children were evaluated of whom 134 (57.8%) were boys and 98 (42.2%) were girls. The Movement Assessment Battery for Children-2 (MABC-2) was used to collect data. The data show a main effect on gender factor; there was a main effect in total score of manual dexterity (MD; p = 0.010), in total score of balance (Bal; p < 0.001), in total test score (TTS; p < 0.001), and in total percentile score (TPS, p < 0.001). In the semester of birth factor, there were differences in aiming and catching (A&C, p < 0.001), in Bal (p = 0.029) and in total percentile score (TPS, p = 0.010). Girls perform better in MD, Bal, TTS, and TPS than boys. Preschool children born in the first semester obtain, in general, a higher percentage and a higher percentile than their peers born in the second one. RAE is present in A&C, Bal, and TPS, with higher scores obtained by preschool children born in the first semester compared to those born in the second one.