Construction of eukaryotic expression vector of human MCHR2 and establishment of stably transfected cell line

AIM: To construct the eukaryotic expression vector of human melanin-concentrating hormone receptor 2 (MCHR2) and stably transfect HEK293 cells with it. METHODS: The full-length MCHR2 cDNA fragment was amplified by PCR from the human fetal brain cDNA library and was inserted into eukaryotic expression vector pcDNA3.1(+). After identification of restriction digestion and PCR, the recombinant plasmid was transfected into HEK293 cells by lipofectamine. After screening culture by G418, a stably-transfected cell line was established, and the transcription and expression of the MCHR2 gene were identified by RT-PCR, Western blot and immunofluorescence assay. RESULTS:The eukaryotic expression vector pcDNA3.1-MCHR2 was successfully constructed and the MCHR2 gene was transfected stably into HEK293 cells. A stably-transfected cell line was establishedand the MCHR2 gene was expressed successfully. CONCLUSION: The establishment of the stably-transfected cell line and the expression of the target gene provide a solid experimental foundation for further studies on the function of the MCHR2 gene.