FIG. 9.
Epo-enhanced activation of βmaj-globin gene expression in FDCW2ER372-pXG1-pEG1 cells. FDCW2ER372-pXG1-pEG1 cells expressing the minimal Epo receptor form ER372 and increased levels of GATA-1 were prepared via stable electrotransfection and maintained in IL-3. Cultures then were subdivided and maintained in parallel in either IL-3 (as WEHI-3 CM) or Epo (25 U/ml). At the indicated intervals of culture (weeks 1, 2, 3, and 4), total RNA was isolated and levels of GATA-1, EKLF and βmaj-globin transcripts were analyzed by Northern blotting. As negative and positive controls, RNA from parental FDCW2ER372 and erythroleukemic SKT6 cells were used, respectively. Hybridizations were performed sequentially with a single blot, and equivalence in loading was confirmed by hybridization to a GAPDH probe.