目的：构建逆转录病毒介导的诱导型一氧化氮合酶（ｉＮＯＳ）基因转移载体．方法：应用ＤＮＡ重组和ＰＣＲ扩增技术．结果：通过二步克隆，从质粒ｐＫＳｉＮＯＳ分离编码巨噬细胞ｉＮＯＳ的全长ｃＤＮＡ，亚克隆于中间载体ｐＳＰ７２，调整插入片段二端酶切位点；进而构建含ｉＮＯＳ基因、巨细胞病毒启动子和ｎｅｏ抗性基因的逆转录病毒载体ｐＬＮＣＸｉＮＯＳ．限制性酶切分析和ＰＣＲ鉴定证实，重组体ｉＮＯＳ插入片段的大小和方向正确．结论：获得含ｉＮＯＳ基因逆转录病毒表达载体，为建立ｉＮＯＳ基因修饰的神经细胞模型，研究ＮＯ／ＮＯＳ在阿片耐受依赖中的作用奠定基础． Objective: To construct a retrovirus-mediated inducible nitric oxide synthase (iNOS) gene transfer vector. Methods: DNA recombination and PCR amplification were used. Results: The full-length cDNA of iNOS encoding macrophage cells was isolated from plasmid pKSiNOS by two-step cloning and subcloned into pSP72 vector to adjust the restriction endonuclease site of the inserted fragment. Then the recombinant plasmid containing iNOS gene, cytomegalovirus promoter and neo resistance gene retrovirus vector pLNCXiNOS. Restriction analysis and PCR analysis confirmed that the size and orientation of the recombinant iNOS insert was correct. CONCLUSION: Obtaining retroviral expression vector containing iNOS gene can lay a foundation for the establishment of iNOS gene modified neural cell model and study the role of NO / NOS in opioid dependence.