Abstract: The amphioxus distributes along the east coast of China from Xiamen waters to Bohai Sea and Japanese waters. It is commonly accepted that amphioxus in both Bohai Sea and Japanese waters belong to Branchiostoma belcheri tsingtauense, a subspecies of B. belcheri, according to the previous survey on the morphological characteristics. Nevertheless, the genetic distance between Xiamen amphioxus and Qingdao amphioxus averages 18.8% and 20.3% based on partial sequence analysis of COI and Cyt b gene, which is much higher than that is observed in other intraspecific variations in the present paper. The values between Japanese amphioxus and Qingdao amphioxus are only 1.3% and 0.9% based on partial sequence analysis of COI and Cyt b gene, which indicates both of them should belong to the same species. Above all, our results support the viewpoint that amphioxus in Qingdao and Xiamen might belong to different species and the original subspecies B. belcheri tsingtauense should be renamed as a separate species and its new scientific name should be B. tsingtauense.