Objective:To construct the lentiviral vector carrying nNOS(AA1-133)and detect its expression and function. Methods:Mouse nNOS(AA1-133)was amplified by RT-PCR and then eukaryotic expression vector of pIRES2-EGFP/nNOS(AA1-133)was constructed. After DNA sequence analysis,nNOS(AA1-133)was cloned into lentiviral vector pGC-FU to construct recombinant vector pGC-FU/nNOS(AA1-133). pGC-FU/nNOS(AA1-133)was transfected into 293T cells by Lipofectamine 2000 mediation to package lentiviral particles. The particles were transfected into primary neurons,and the expression and function of nNOS(AA1-133)were detected. Results:The coding sequence of mouse nNOS(AA1-133)was successfully amplified and the analysis of DNA sequence approved that the recombinant lentiviral vector contained nNOS(AA1-133). nNOS(AA1-133)expressed and combined with PSD95 in the infected neurons. Conclusion:The lentiviral vector of pGC-FU/nNOS(AA1-133)was constructed successfully and the infected neurons could express nNOS(AA1-133).