Figure 2.

Alpharetroviral vectors containing a myeloproliferative sarcoma virus (MPSV) promoter and the modified feline endogenous retrovirus envelope glycoprotein RD114/TR deliver enhanced gene transfer into cord blood (CB)-derived CD34+ hematopoietic stem cells. (a) Self-inactivating (SIN) alpharetroviral vectors containing either an MPSV or an elongation-factor-1-short-form (EFS) promoter combined with an enhanced green fluorescent protein (EGFP) reporter gene were generated. Indicated are unique 5 (U5), repeat (R), and SIN unique 3 (ΔU3) regions, long terminal repeat (LTR), packaging signal (ψ), woodchuck posttranscriptional regulatory element (PRE) and direct repeat element (DR). (b) MPSV- and EFS-driven alpharetroviral vectors were pseudotyped with either vesicular stomatitis virus glycoprotein (VSVG) or RD114/TR and used to transduce CD34+ cells using an equal multiplicity of infection (MOI) of 100. (c,d) Transduction efficiency and mean fluorescence intensity (MFI) of EGFP were determined 6 days after transduction. Representative results of two independent experiments are shown (n = 3). (e) CB CD34+ cells were transduced with an increasing MOI using alpharetroviral MPSV- or EFS-driven vectors with the EGFP gene. After 6 days, transduction efficiency was determined by flow cytometry. Mean vector copy number (VCN) was assessed using quantitative real-time PCR for detection of PRE within the vector.