The γ/β-globin switch is recapitulated in the one-phase liquid culture system. Cells grown in the one-phase system were harvested every 2-3 days and analyzed as follows (see Methods). A) Cell morphology was determined by Giemsa stain and different cell types classified based on standard morphologic features. B) RT-qPCR analysis was performed to determine changes in γ-globin and β-globin mRNA levels during erythroid maturation. C) RT-qPCR was performed with Gγ and Aγ specific primers to determine the Gγ:Aγ mRNA ratio during erythropoiesis. Aγ-globin expression was normalized to one (white bars) and the relative changes in Gγ-globin mRNA are shown in the black bars. D) Progenitors were induced on day 11 or day 28 with 50 μM hemin (HE) for 72 hrs and then RT-qPCR was completed.
Li et al. BMC Genomics 2012 13:153 doi:10.1186/1471-2164-13-153