Genetic reshuffling reconstitutes functional expression cassettes in retroviral vectors

Background A major prerequisite for the design of retroviral vectors encoding cell toxic or harmful genes is the possibility to tightly control gene expression, thus limiting activity to the relevant target cells and protecting the packaging cell used for production of recombinant viral particles. Methods In the present study a system was developed in which genetic reshuffling during the retroviral life cycle is exploited, allowing reconstitution of functional expression cassettes from separate elements exclusively in transduced target cells. For construction of these murine leukaemia virus (MLV)-based reconstituting viral vectors (ReCon), a promoterless inverted enhanced green fluorescent protein (EGFP) reporter gene cassette was inserted in place of the U3 region of the 3k LTR. Subsequently, the human ubiquitin promoter was inserted in the inverse orientation into the R/U5 border of the 5k LTR of the vector.