Establishment of a spleen cell line from large yellow croaker Pseudosciaena crocea and its primitive application in foreign gene transfection

Abstract

A large yellow croaker, Pseudosciaena crocea, spleen (LYCS) cell line was established and the feasibility of using it for foreign gene transfection was evaluaed in this study. Primary culture of LYCS cells was initiated from spleen tissue pieces, which were cultured at 25°C in Dulbecco’s modiced Eagle medium/F12 medium (DMEM/F12, 1:1) (pH7.2), supplemented with 20% fetal bovine serum, carboxymethyl chitosan, chondroitin sulfate, basic fibroblast growth factor (bFGF) and insulin-like growth factor-I (IGF-I). The cultured LYCS cells, in fibroblast shape, proliferated to 100% confluency 20 days later. Chromosome analyses indicated that the LYCS cells exhibited chromosomal aneuploidy with a modal chromosome number of 48 which displayed the normal diploid karyotype of P. crocea (6m+6sm+36t, NF=60). A LYCS cell line, with a population doubling time of 48.7 h at passage 60, has been established and subcultured to passage 70. Transgenic feasibility test demonstrated that positive green fluorescence protein (GFP) expression was observed in LYCS cells after pcDNA3.1-GFP plasmid transfection. In conclusion, a continuous foreign gene transfection feasible LYCS cell line has been established successfully. The cell line might serve as a valuable tool for studies of transgenic breeding and has potential applications for different kinds of cytotechnological studies.