Establishment and characterization of a new cell line from the kidney of spotted halibut Verasper variegates

Abstract

A cell line, SHK, was derived from the kidney of spotted halibut Verasper variegates. The cell line was subcultured more than 40 passages in minimum essential medium (MEM) supplemented with fetal bovine serum (FBS) and 10 ng ml−1 basic fibroblast growth factor (bFGF). Cell morphology from primary culture and subculture was observed continuously by microscopy. The SHK cell line consisted predominantly of fibroblast-like cells. The cell line was able to grow between 20°C and 30°C with the optimum growth at 24°C and with a reduced growth between 12°C and 20°C. The growth rate of the cells increased as the proportion of FBS increased from 10% to 20% at 28°C with optimum growth at the concentration of 20%. The doubling time of the cells was determined to be 44.8 h. Chromosome analysis revealed that 52% of the SHK cells maintained a normal diploid chromosome number (2n=46). The cells were successfully transfected with green fluorescent protein (GFP) reporter plasmids and the expression of GFP gene in the cells indicated the possible utility of the cells in gene expression studies. The cells were infected by lymphosystis disease virus (LCDV) and found to be susceptible to the virus in cytopathic effect (CPE) observation. The infection was confirmed by PCR and electron microscopy experiments, which proved the existence of the viral particles in the cytoplasm of the virus-infected cells.