Three mitochondrial (mt) genes were sequenced for two Atlantic lancelet species, Branchiostoma lanceolatum and B. floridae, to examine a serious discrepancy among previously published results of molecular studies: substantial sequence difference in a nuclear gene vs. virtual identity in the mt genome sequence. The results revealed that three mt genes of B. lanceolatum, collected from Helgoland in the North Sea and Naples in the Mediterranean, were quite diverged from those of B. floridae, collected from Tampa Bay, Florida. Therefore, the previously recognized identity in the mt genome between the two species is attributable to misidentification of materials used. To correct this misleading information, the complete mtDNA sequence of B. lanceolatum was determined for an individual from Helgoland.