We used functional magnetic resonance imaging to investigate the cortical mechanisms contributing to the acquisition and performance of a complex, bimanual motor sequence. To that aim, five subjects were trained on a difficult, asymmetrical finger opposition task. Their performance rate almost doubled in the course of training and approached the performance rate in an untrained, symmetrical finger opposition task. Before training, performance of the asymmetrical sequence was associated with activity in M1, premotor cortex, supplementary motor cortex, and parietal cortex. After training, performance of the asymmetrical sequence was associated mainly with activity in M1, and little activity outside M1 remained. The latter pattern of cortical activation resembled that observed during the execution of symmetrical sequences, which was unaffected by practice with the asymmetrical sequence. The activation pattern obtained with the symmetrical bimanual sequence was indistinguishable from the combined activation measured in contralateral hemispheres during unimanual control sequences. The data indicate that cortical regions previously implicated in the acquisition of difficult unimanual motor sequences also contribute to the acquisition of asymmetrical bimanual sequences. We found no evidence for an expansion of activity in M1 after acquisition of the asymmetrical sequence (while this has been reported after acquisition of unimanual sequences). In the context of existing literature, the data suggest that the acquisition of unimanual and bimanual motor sequences may rely on similar cortical mechanisms, but that the formation of long-term, procedural memories for the two types of sequences might at least in part depend on different mechanisms.