Objective To establish a stable transfection cell line of iRhom2 and its mutant through recombinant lentivirus infection. Methods The full-length gene of iRhom2 and its mutant were cloned into the lentivirus vector Lenti-OE-Flag, and got recombinant lentiviral vector of Lenti-OE-iRhom2 and Lenti-OE-iRhom2mut. The constructed recombinant lentivirus vectors were transfected into HEK-293T packaging cells to obtain the recombinant virus. Vero cells were infected with recombinant virus. The stable expressing cell lines were obtained by pressure screening with puromycin. Results The recombinant lentivirus vectors were constructed and the recombinant virus was obtained. The stable expressing cell lines were obtained using virus infection and the protein expression was testified with Western blotting. Conclusions Stable iRhom2-expressing Vero cell line and its mutant are achieved by recombinant lentivirus infection. It paves the way for future study on biological functions and mechanism of iRhom2.