We recently reported on an hematopoietic stem cell (HSC) gene therapy approach. It involves the subcutaneous injections of G-CSF/AMD3100 to mobilize HSCs from the bone marrow into the peripheral blood stream and the intravenous injection of an integrating helper-dependent adenovirus vector system. HSCs transduced in the periphery homed back to the bone marrow， where they persisted long-term. However， high transgene marking rates found in primitive bone marrow HSCs were not reflected in peripheral blood cells. Here， we tested small-molecule drugs to achieve selective mobilization and transduction of HSCs. We found more efficient GFP marking in bone marrow HSCs but no increased marking in the peripheral blood cells. We then used an HSC chemo-selection based on a mutant of the O-methylguanine-DNA methyltransferase (mgmt) gene that confers resistance to O-BG/BCNU and should give stably transduced HSCs a proliferation stimulus and allow for the selective survival and expansion of progeny cells. Short-term exposure of G-CSF/AMD3100-mobilized， -transduced mice to relatively low selection drug doses resulted in stable GFP expression in up to 80% of peripheral blood cells. Overall， the further improvement of our HSC transduction approach creates the basis for a simpler HSC gene therapy.