ZNF648 knockdown impedes megakaryocyte differentiation. Isolated adult peripheral blood CD34+ cells were transferred to megakaryocyte differentiation medium and transduced with scrambled (Scr) short hairpin RNA (shRNA) as control or ZNF648 shRNA followed by puromycin selection. Untransduced cells served as a further control. (A) Schematic of experimental design and expression profile of CD34, CD41, CD61 and CD42b during normal megakaryopoiesis, (B) expression of membrane markers CD34, CD41 (platelet glycoprotein IIb), CD61 (platelet glycoproteins IIIa) and CD42b (GPIb ) analysed by flow cytometry with antibodies CD41- PE, CD61-APC (both from Biolegend), CD34-VB BD and CD42b-PE BD (both from Pharming). Data was acquired with a MacsQuant VYB Analyser using a plate reader, n=6. (C) Proportion of CD41/CD61 and CD61/CD42b positive cells on day 14 of culture, n=6 ± standard error of the mean. (D) Ploidy status of cells at day 14 of culture, n=3. (E) Morphology of cells at day 14 of culture stained with May-Grünwald-Giemsa. Upper panel 100x magnification, lower panel 400x magnification. Images representative of three independent cultures. *P<0.05, **P<0.01, ***P<0.001, Student’s t-test.