The stable introduction of a functional gene into autologous stem cells is a potentially powerful approach to treat a number of inherited or acquired diseases. One challenge facing this approach is to express adequate levels of the therapeutic transgene in a regulated and sustained fashion, eventually restricting expression to a single lineage developing from the transduced stem cells. Until now, low-level expression, position effects, and transcriptional silencing have hampered the effectiveness of retroviral-mediated gene transfer. In an effort to overcome these obstacles, we have systematically investigated vectors encoding the human beta-globin gene linked to selected combinations of proximal and distal genetic regulatory elements. Our results demonstrate that with thoughtful vector design one can successfully express long-term, therapeutic levels of virally encoded human beta-globin in the erythroid progeny of hematopoietic stem cells.