Cotransduction with MGMT and Ubiquitous or Erythroid-Specific GFP Lentiviruses Allows Enrichment of Dual-Positive Hematopoietic Progenitor Cells In Vivo
Cotransduction for selection and lineage-specific expression in vivo. Donor bone marrow cells were cotransduced with MGMT and RRL-GATA-GFP viruses for 12 hours prior to transplant into lethally irradiated recipients. PB AGT+ and TER119+-GFP+ expression percentages were taken at three week intervals, two days prior to each treatment with 30 mg/kg BG and 10 mg/kg BCNU. (a) AGT+ and TER119+-GFP+ percentages in lethally irradiated mice transplanted with SKL cells cotransduced with an MGMT MOI of 5 and an RRL-GATA-GFP MOI of 15. (b) Flow cytometric analysis of GFP expression in TER119+ bone marrow erythroblasts from animals 1 through 6 in Panel (a) (hatched bars). The percentage positive in each quadrant is listed, as well as the percentage of GFP expression restricted to TER119+ erythroblasts (boxed). (c) Expression of RRL-GATA-GFP is restricted to TER119+ cells. (d) Analysis of MGMT (“AGT”, closed circles) and GFP (open squares) copy numbers per diploid genome in BM CFU obtained from Panel (a) animals 1 through 6 (hatched bars).
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