PRODUCTION OF HIGH-TITER HELPER-FREE RETROVIRUSES BY TRANSIENT TRANSFECTION

The generation of high-titer, helper-free retroviruses by transient tr ansfection has been achieved by using the highly transfectable 293T ce ll line into which are stably introduced constructs that express retro viral packaging functions. The resulting ecotropic virus packaging cel l line BOSC 23 produces infectious retrovirus at >10(6) infectious uni ts/ml of supernatant within 72 hr after CaPO4-mediated transfection. A stringent assay for replication-competent virus showed that no helper virus was present. The system can produce high titers of retroviral v ectors expressing genes that are extremely difficult to propagate at h igh titer in stable producer lines. This method should facilitate and extend the use of helper-free retroviral gene transfer, as well as be useful for gene therapy.