Recent reports show that MIDI keyboard playing can improve upper-limb functions in neurorehabilitation, but little attention has been paid to this type of intervention with stroke patients in the subacute stage. The purpose of this study was to investigate the effects of keyboard-playing intervention that targeted hand dexterity of subacute stroke patients. A total of 17 subacute stroke patients participated in twelve 30-minute sessions over six weeks. The intervention included repetitive individuated, sequential, and simultaneous key-pressing to increase independent finger movements and strength. Of the four hand-function tests administered in this study, tip pinch in hand grip and pinch strength test and Box and Block Test (BBT) results showed the most significant improvement, indicating the largest effect sizes (0.785 for hand grip and pinch strength test and 0.840 for BBT). Better hand dexterity for subacute stroke patients may be achieved through repetitive key-pressing motions during keyboard playing by exercising the intrinsic and extrinsic flexor muscles primarily involved in the key-pressing motion. Overall, this study supports keyboard playing as a promising therapeutic intervention in subacute stroke rehabilitation.