In this study, we established and characterized a cell line derived from the kidney of black carp (Mylopharyngodon piceus), which is an important freshwater aquaculture species. The cell line was designated as MPK and subcultured for more than 70 passages in DMEM medium containing 10% fetal bovine serum (FBS) at 28°C. MPK had a modal diploid chromosome number of 48. Moreover, a transient MPK transfection efficiency was up to 18% using a green fluorescent protein plasmid by a modified electroporation. In addition, the MPK cells showed susceptibility to spring viremia of carp virus (SVCV), as demonstrated by the presence of severe cytopathic effects (CPEs) and increased viral RNA. Unexpectedly, the MPK cells expressed pluripotency-associated genes such as nanog, oct4 and vasa, indicating that these are possibly adult stem cells. Taken together, we have established a stable cell line from kidney that may potentially be utilized as an in vitro platform for genetic modifications and host-pathogen analysis in black carp.