Fli-1 gene deletion decreases the megakaryocytic differentiation potential of MEPs at the benefit of erythrocytic differentiation. (A-C) Histograms showing the number of different classes of colonies generated in semisolid medium containing a complete cocktail of cytokines (IL3, IL11, TPO, EPO, SCF, GM-CSF, Flt3-l) by 100 sorted CMPs (A), GMPs (B), or MEPs (C; means and SDs from 5 different couples of injected fli1^fl/fli1^Δ Cre⁺ and Cre⁻ mice). Note that the loss of virtually all identifiable megakaryocytic and mixed colonies in the progeny of Cre⁺ MEPs in C occurred without significant decrease of overall clonogenicity. (D) PCR analyses of Fli-1 gene deletion in single erythrocytic (E) or megakaryocytic (Mk) colonies derived from Cre⁺ or Cre⁻ MEPs purified from Fli1^Δ/Fli1^fl mice. The top gel (lanes 1-7) has been generated using primers allowing the detection of undeleted allele Fli1^fl only, whereas the bottom gel (lanes 8-14) has been generated using primers allowing simultaneous detection of deleted Fli1^Δ and undeleted Fli1^fl alleles. Lanes 1, 8, and 9 correspond to tail DNA from uninjected control mice. (E) Single-sorted bone marrow MEPs from injected Cre⁻ or Cre⁺ mice were seeded at 1 cell per well in liquid medium containing complete cocktail of cytokines (IL3, IL11, TPO, EPO, SCF, GM-CSF, Flt3-l), and the cell composition of each well was individually scored under light microscopy after 1 week of culture. Histogram shows Cre⁺/Cre⁻ ratios for each class of wells containing either pure erythrocytic colony (large number of small cells and no identifiable large mature megakaryocyte), mixed colony (large number of small cells with clearly identifiable megakaryocytes), and pure megakaryocytic colony containing the indicated number of megakaryocytes (typical light microscope views of colonies are given in supplemental Figure 2). Means and SDs from analyses of 6 different couples of Cre⁺ and Cre⁻ mice (3 couples of Fli1^fl/Fli^fl and 3 couples of fli1^fl/fli1^Δ genotypes). All Cre⁺/Cre⁻ ratios were significantly different from 1. NS indicates nonsignificant differences between different classes of megakaryocytic colonies (P > .05) in paired Student t test.