We investigated the formation kinetics of a single monolayer nanotube from bolaamphiphiles (consisting of a sugar residue, an alkyl chain, and an amino group) in solution. In this bolaamphiphile, a transition from a monomerically dispersed state to the nanotube takes place by changing the solvent condition. This transition was induced by fast mixing with a stopped-flow apparatus. From just after the mixing, this transition process was monitored in situ by time-resolved small-angle X-ray scattering. In this manner, we were able to derive the direct structural information as a function of time during the nanotube formation. The results revealed that disklike aggregates initially formed, which then grew and closed to produce a tubular structure.