Biological invasions are among the major global causes of biodiversity loss. The pet trade is the main pathway by which reptiles are introduced into new environments. After the EU import ban on the pond slider (Trachemys scripta), other species of pet-traded freshwater turtles are more often reported from the wild. Here we report the first and genetically verified record of Chinese soft-shelled turtle (Pelodiscus sinensis) from the eastern part of the Czech Republic. The genus Pelodiscus has been reported from the Czech Republic and neighbouring countries; however, only as a visual record, which is insufficient for identification due to the complexity of the taxonomic status of this species. We physically captured the turtle and used genetic data (sequences of 16S rRNA and cytochrome b) to confirm the species affiliation as P. sinsensis. The studied individual represents a lineage of P. sinensis that is widely bred on farms with a native range in North Vietnam, South and south-east China.