Abstract: Objective To construct recombinant over expression vector of Homo sapiens proenkephalin (PENK) gene and explore the function of PENK gene. Methods Fragment containing PENK ORF gene was inserted into vector plasmid HIV, then the recombinant over was confirmed by enzyme digestion and sequencing. Lentivirus containing the recombinant over expression vector was produced by virus packaging with 293Ta cell，and then the lentivirus was transfected into HT1080 cell and the virus titer was estimated. The PC12 were tansfected with resulting lentivirus and un-transfected PC12 cells as control. The images of the PC12 cells were captured at 48 h post-transfection and the number of cells was also evaluated; the changes of PENK mRNA in transfection and control group were measured with RT-PCR. Results The constructed PENK ORF recombinant over expression vector was confirmed by enzyme digestion and sequencing. The number of PC12 cells in transfection and control group at 48 h post-transfection was 127.93±2.48 and 88.60±2.55 respectively, and the statistical difference between them was observed (P<0.01). Conclusion Recombinant over expression vector of PENK gene was successfully constructed and the PENK gene can promote the growth of PC12.