A transient three-plasmid expression system for the production of high titer retroviral vectors

摘要

We have constructed a series of MLV-based retroviral vectors and packaging components expressed from the CMV promoter and carried on plasmids containing SV40 origins of replication. These two features greatly enhanced retroviral gene expression when introduced into cell lines carrying the SV40 large T antigen. The two packaging components, gag-pol and env, were placed on separate plasmids to reduce helper virus formation. Using a highly transfectable human cell line and sodium butyrate to further increase expression of each component, we achieved helper-free viral stocks of ～107 Infectious units/ml by 48 h after transient co-transfectlon with the three plasmid components. This system can be used both for the generation of high titer retroviral stocks for transductlon and for the rapid screening of a large number of MLV gag-pol or env mutants.