There are only a few cell lines of Siniperca chuatsi (mandarin fish) that could be widely used in scientific research. For mandarin fish, cell culture technique is an important aspect of the quality control program for the National Wild Fish Health Survey. Healthy, sensitive and mycoplasma-free cells are essential for detection of fish viruses in free-ranging fish populations. S. chuatsi is native to China and an important commercial species. In addition, the transfection efficiency is quite low in most fish cell lines. That is an obstacle in the basic research on fishes. In this study, a continuous cell (MFE) culture derived from the mandarin fish embryo was developed and has been subcultured over 20 passages in DMEM cell culture medium. MFE consists predominantly of epithelial-like cells and grows well in DMEM supplemented with 20% fetal bovine serum. Cell proliferation was assessed by MTT assay. The absorbance of the sample was read directly in the wells at an optimal wavelength of 570 nm. The results showed that MFE cell experienced proliferation, decrease and stable phases during 96 h. Meanwhile, we used GFP-expressing retrovirus produced by HEK293T cells to test the transfection efficiency in MFE cell line. The result showed that the transfection efficiency reached 20%±5% without affecting the growth of these cells. Subsequently, the expression of three genes of Irf1, Irf2 and Irf7 was examined in MFE cell line by qRT-PCR. The results indicated that Irf1 is the highest one, and all increased to 3.5, 2.3 and 2.1 folds after poly I:C stimulation. The results of this study showed that MFE is the first cell line originated from embryo of mandarin fish. It could be used as a tool for gene function research and genetic modification.