A, genomic DNAs from a polyclonal population of BaF3.EpoR.JAK2V617F cells, containing 1% of JAK2V617F mutant expressing cells were analyzed at day 0 and day 7 after treatment with either DMSO (control) or ruxolitinib (300nM) for the presence of E985K and Y931C substitution (arrows indicate position of corresponding base substitution). Partial chromatograms of the forward strand are shown. B, expression of STAT5, phospho-Y694 STAT5 and β-actin was determined by immunoblotting in BaF3.EpoR cells expressing either JAK2V617F (control) and cells containing the additional E985K or Y931C mutation. Cells were either treated with DMSO or 300 nM ruxolitinib. C, relative changes in EC50 values were determined in response to ruxolitinib for BaF3.EpoR.JAK2V617F.M929I cells, relative to cells expressing native JAK2V617F.