Establishment And Application Of A Cell Line Derived From Fin Of Squaliobarbus Curriculus

Cell culture technology has been widely applied used in biological research, based on more GCRV resistant of Squaliobarbus curriculus than grass carp,provide new cell resouces for breeding of disease resistant vavieties of Grass carp. Established and applicated of a cell line derived from fin of Squaliobarbus curriculus, obtained the following results:1.A cell line desagnated SCF, has been established and characterized from fin of Squaliobarbus curriculus,using tissue explant techniques. After the incubation of 7-8 days, the cells were observed under the microscope, and the cells were moved out from the periphery of the tissue. After the primary culture for about 14 days, the tissue mass was surrounded by a halo. SCF cell line belongs to fiber-like cells, and has been successfully subculture over by the method of trypsin digestion。2.The studies on the optimal cultureconditions of SCF cells include the optimal serum concentration,sutible culture medium. The results show that the optimal concentration of serum was 20%.The optimal medium was L-15 medium.After recovery, SCF cells exhibit a viability of 82.5%.Chromosome analysis of this cell line revealed that at its 25th passage the diploid chromasome number was 2n=48.In order to further prove that the cells derived from Squaliobarbus curriculus,a pair of specific primers of 18SrRNA of Squaliobarbus curriculus was synthesized and the 18SrRNA of Squaliobarbus curriculus was amplified.Seqence analysis showed that the amplified fragment with a length 185bp showed 97% identity with the 18SrRNA gene of Squaliobarbus curriculus.3.The results of virus sensitivity test indicated that SCF cell line is susceptible to the infection of GCRV and GISV.A haracteristic viral-induced cytopathic effect(CPE) was observed and viral titer(TCID50/ML) respectively reached about 105.33and 101.48. The pEGFP-N1 plasmid transfected into Squaliobarbus curriculus fin cells and fluorescence microscopy of EGFP expressed green fluorescence, indicating that the cells can be used for the expression of exogenous gene.The establishment of SCF cell line ont only enrichs the fish cell line resources,but also provided an important experimental material for the studies on immune related of GCRV.