Bioluminescence imaging reveals the long-term dynamics of individual gene expression in a single cell. However, methods for simultaneously imaging multiple gene expression patterns have been unknown to date. Here, we constructed a dual-path optical luminescence imaging system using a two-color reporter system and could simultaneously track two gene expression patterns for several days in a single cell. Address correspondence to Yoshihiro Ohmiya, Regenerative Medicine/Tissue Engineering Division, Research Center for Cooperative Projects, Hokkaido University Graduate School of Medicine, Kita-15, Nishi-7, Kita-Ku, Sapporo 060-8638, Japan. e-mail: