Construction of an inducible nitric-oxide synthase gene transferring vector mediated by retrovirus

AIM: To construct an inducible nitric-oxide synthase (iNOS) gene transferring vector mediated by retrovirus. METHODS: Recombinant DNA and polymerase chain reaction (PCR) amplification techniques were used. RESULTS: With 2 steps of molecular cloning, the full-length cDNA encoding macrophage iNOS was isolated from plasmid pKSiNOS and subcloned into intermediate vector pSP72, adjusting the restriction enzyme sites in both 5'- and 3'-flanking ends of insert fragment. The retroviral vector pLNCXiNOS which contains iNOS coding region, cytomegalovirus promoter and neomycin resistance (neo(r)) gene was further constructed. The authenticity of insertion size and orientation of iNOS sequence was verified by restriction mapping and PCR analysis with iNOS gene-specific primers. CONCLUSION: Retroviral expression vector carrying iNOS fragment is obtained, which provides a material to establish a model of iNOS gene-modified neurons.