Spontaneous division of lipid bilayer vesicles was observed for giant vesicles formed by a phospholipid mixture containing a nonbilayer-forming lipid, i.e., a nonionic peptide lipid with a quinoyl head moiety. This synthetic cell division involved dynamic sequential processes, including phase separation of mixed lipids in the vesicles, vesicle deformation from a spherical to an elliptical shape, and subsequently fission.