Figure 1.

Timing of acquisition of DNMT3A mutations. Single cell-derived hematopoietic erythroid colonies (BFU-E) were grown in vitro from peripheral blood-derived mononuclear cells and individually genotyped for mutations using Sanger sequencing. Plots in (A), (B), (C) and (D) show colony genotyping results for mutations in DNMT3A and JAK2, MPL or CALR for each patient and the order of mutation acquisition. Within each plot, each dot represents a single colony and its quadrant placement shows the corresponding genotype of DNMT3A (vertical axis) and JAK2/MPL/CALR (horizontal axis). Solid red arrows within quadrants show the confirmed path of clonal evolution. Wt, wild-type; het, heterozygous mutation; hom, homozygous mutation; PV, polycythemia vera; ET, essential thrombocythemia; PMF, primary myelofibrosis; PPV-MF, post-PV myelofibrosis (A) DNMT3A-first patients: four patients in whom mutated DNMT3A occurred prior to the acquisition of JAK2^V617F. (B) Biclonal patients: three patients in whom DNMT3A^mut and JAK2^V617F were in separate clones. (C) JAK2/MPL-first patients: three patients in whom mutated JAK2 or MPL occurred prior to the acquisition of mutated DNMT3A. (D) Order unknown: three patients in whom the order of mutation acquisition of DNMT3A and JAK2/MPL/CALR could not be delineated as only wild-type colonies and/or double mutant colonies were detected.