Abstract: [Objective] To establish immortalized ovine ruminal fibroblast cell line and to provide stable in vitro cell model for basic research and animal virus research. [Method] Ovine ruminal fibroblast cells（ORFCs） were cultured by using tissue fragment clinging method. pCI-neo-hTERT plasmid with hTERT gene was transferred into ORFCs using PEI reagents. The positive clone cells were screened by G418. The transcription and expression level of hTERT gene in 15^th and 35^th generation positive clone cells were detected by RT-PCR and Western Blot. The growth curves of primary cells and 35^th generation positive clone cells were drawn with blood cell count method to assess the proliferation ability. [Result] The hTERT gene was successfully transferred into ORFCs and stably expressed. The proliferation ability of positive clone cells was significantly higher than that of primary ORFCs. [Conclusion] Immortalized ovine ruminal fibroblast cell line is successfully established.

Key words: ovine ruminal fibroblast cell line, immortalization, human telomerase reverse transcriptase