Establishment Of Vitro Culture System And Biological Characteristics For Three Kinds Of Lynx Fibroblast Cells

To study the culture system in vitro and biological characteristic on three kinds of lynx cells and do further development about lynx cell genomics with providing a reliable material platform for project of wildlife protection, the cells from the tissue of heart, embryo body and testis of lynx were got by tissue culture in this experiment, and a-MEM medium was used for the primary culture and serial subcultivation, conventional methods for cryopreservation and recovery were performed in order to establish culture system in vitro. Then morphology observation, growth curves, microbial detection, karyotype analysis and fluorescent protein plasmid transfection and expresstion were carried out for their biological characteristic analysis. The results showed that there were some differences between different tissue cells in lynx, ranging from morphosis to growth cycle. During primary culture, the growth rate of different tissue cells showed difFerences.However there were not significant differences among serial subcultivation. Compared with before frozen, the survival rate of the cells after thawed was lower than before. But both were maintained at90%. The three kinds of lynx cell all showed a growth curve "S" type, every generation has experienced growth latency, exponential phase of growth and stagnation period, consistented with in vitro cell growth pattern. Detection for bacteria, fungi, virus and mycoplasma were negative.The further karyotype analysis indicated that the number of lynx cell chromosomes was38, including18pairs of autosomes and one pair of sex chromosomes. The result showed diploid cells were dominant of88%, then genetic characteristics of cell culture is stable. Liposome-mediated with red fluorescent protein reporter plasmid transfection showed that three tissue fibroblasts of lynx were transfected, and the transformation efficiency were58%,51%and46%respectively. The result meaned the cell cultured in vitro could transfer and express foreign genes effectively. Conclusion:we has successfully established the culture system of three kinds of tissue fibroblast cells in vitro, and then genetic resource of lynx was conserved in cell level. These results provided experimental material and the basis for further research in the field. The study also will be a useful reference for the establishment of lynx cell line, the better use of genetic resources and the protection of endangered species of precious wild animals.