Abstract: Transgenic efficiency in mammal cell lines is usually higher but relatively lower in the fish cell lines. Therefore, establishment of any fish cell lines with high transfection efficiency is needed, ideally in the fish model such as medaka (Oryzias latipes). In the present study, we derive a new cell line from medaka muscle termed as OLM (Oryzias latipes muscle), which was maintained in DMEM/HEPES supplemented with 10% bovine serum at 28℃. Up to now, OLM cells have been subcultured over 88 passages and mainly exhibited a fibroblast-like morphology. By chromosomal analysis, OLM has a modal diploid chromosome number of 48. Moreover, transient transfection efficiency by liposome of OLM cell line is up to 40%, which is much higher than that of the common fish cell lines as tested. In addition, the stable OLMs expressing exogenic non-coding RNA or gene are established. Unexpectedly, the OLM is resistant to the fish virus such as Spring viraemia of carp viruses (SVCV), Grass carp reovirus (GCRV) and Singapore grouper iridovirus (SGIV). Taken together, we have established a stable muscle cell line from model fish medaka as an in vitro model for transgenic application.