The Ly-6A (Sca-1) GFP transgene is expressed in all adult mouse hematopoietic stem cells

The Sca-1 cell surface glycoprotein is used routinely as a marker of adult hematopoietic stem cells (HSCs), allowing a >100-fold enrichment of these rare cells from the bone marrow of the adult mouse. The Sca-1 protein is encoded by the Ly-6A/E gene, a small 4-exon gene that is tightly controlled in its expression in HSCs and several hematopoietic cell types. For the ability to sort and localize HSCs directly from the mouse, we initiated a transgenic approach in which we created Ly-6A (Sca-1) green fluorescent protein (GFP) transgenic mice. We show here that a 14-kb Ly-6A expression cassette directs the transcription of the GFP marker gene in all functional repopulating HSCs in the adult bone marrow. A >100-fold enrichment of HSCs occurred by sorting for the GFP-expressing cells. Furthermore, as shown by fluorescence-activated cell sorting and histologic analysis of several hematopoietic tissues, the GFP transgene expression pattern generally corresponded to that of Sca-1. Thus, the Ly-6A GFP transgene facilitates the enrichment of HSCs and presents the likelihood of identifying HSCs in situ.