High-efficiency gene transfer into cord blood–derived CD34⁺ cells.

(A) Schematic diagram of FMEV-based retroviral vectors that carry either the bicistronic 2A cassette for coexpression of GFP and HOXB4 (FMEV-HAHOXB4wPRE vector), the GFP marker alone, or the RFP reporter gene alone. All vectors contain the posttranscriptional regulatory element of the woodchuck hepatitis virus (wPRE), which enhances transgene expression. (B) Flow cytometric analysis of reporter gene expression (GFP, RFP) in human CD34⁺ cells 24 hours after the last infection with infectious HOXB4-, GFP-, or RFP-vector particles. Target CD34⁺ cells were prestimulated for 24 hours in cytokine-supplemented, serum-free medium. On 3 consecutive days, the cells were then exposed to RD114-pseudotyped retroviral particles that had been preloaded onto new retronectin-coated plates. The percentage of CD34⁺ cells expressing GFP or RFP is indicated in each dot-blot diagram.