Abstract:  The aim of this study was to construct expression vector containing Lactoferricin gene and interferon-α gene which was transfected into the bovine fetal fibroblasts. The transgenic cells got would be used as the donor cells for SCNT to produce the transgenic bovine that would resist mastitis disease and FMD. The vector with a bovine Lactoferrincin B cassette containing a goat β-casein regulatory sequence, a human interferon-α cassette containing the immediate early promoter of CMV, the EGFP gene as reporter and the neo gene as positive selection marker, was constructed. FuGENE Transfection Reagent and Amaxa Nucleofector were used in transfection. After 48 hours，the cells were selected with G418 and detected by PCR. The expression of interferon-α protein was detected by Western blotting. The transgenic bovine fetal fibroblast cell lines were constructed in which Lactoferricin and interferon-α genes were integrated and the expression of interferon-α protein was detected.