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We developed a selectable marker rendering human cells resistant to Diphtheria Toxin (DT). The marker (DT R ) consists of a primary microRNA sequence engineered to downregulate the ubiquitous DPH2 gene, a key enzyme for the biosynthesis of the DT target diphthamide. DT R  expression in human cells invariably rendered them resistant to DT  in vitro , without altering basal cell growth. DT R -based selection efficiency and stability were comparable to those of established drug-resistance markers. As mice are insensitive to DT, DT R -based selection can be also applied  in vivo . Direct injection of a GFP-DT R  lentiviral vector into human cancer cell-line xenografts and patient-derived tumorgrafts implanted in mice, followed by systemic DT administration, yielded tumors entirely composed of permanently transduced cells and detectable by imaging systems. This approach enabled high-efficiency  in vivo  selection of xenografted human tumor tissues expressing ectopic transgenes, a hitherto unmet need for functional and morphological studies in laboratory animals.

A diphtheria toxin resistance marker for in vitro and in vivo selection of stably transduced human cells