Purpose: The purpose of this study is to use P300 latency to determine whether methods of motor learning in terms of massed and distributed practice can affect motor sequential learning in healthy adults. Methods: Twenty-four healthy subjects participated in this study. They were randomly allocated into three groups: a 10 minute, a 12 hour, and a 24 hour group. In the SRT task, eight numbers were adopted as auditory stimuli. During an experiment, participants were instructed to press the matching key as quickly and accurately as possible when one of the eight numbers was presented randomly. The subjects practiced for three sessions, each of which comprised five blocks of 40 serial reaction time tasks. While they practiced during these three sessions, P300 latency was measured. The data were analyzed using ANCOVA. Results: The P300 latency of Fz, Cz, and Pz decreased in all groups except for the Fz area of the 10 min group. Overall, the P300 latency of the 10 min group showed a smaller decrease compared with the 12 hr and 24 hr groups. Statistically, no significant differences in the Fz and Cz areas were observed among the three groups. The P300 latency in the Pz area of the 10 min group showed a significantly smaller decrease compared with the other groups. Conclusion: These findings suggest that short-term sequential motor training can alter brain functions such as the P300 latency. We also found that better acquisition of a motor skill was obtained with distributed practice of a task than with massed practice.