Although other mutations may predate the acquisition of the JAK2(V617F) mutation, the latter is sufficient to drive the disease phenotype observed in BCR-ABL-negative myeloproliferative neoplasms (MPNs). One of the consequences of JAK2(V617F) is genetic instability that could explain JAK2(V617F)-mediated MPN progression and heterogeneity. Here, we show that JAK2(V617F) induces the accumulation of reactive oxygen species (ROS) in the hematopoietic stem cell compartment of a knock-in (KI) mouse model and in patients with JAK2(V617F) MPNs. JAK2(V617F)-dependent ROS elevation was partly mediated by an AKT-induced decrease in catalase expression and was accompanied by an increased number of 8-oxo-guanines and DNA double-strand breaks (DSBs). Moreover, there was evidence for a mitotic recombination event in mice resulting in loss of heterozygosity of Jak2(V617F). Mice engrafted with 30% of Jak2(V617F) KI bone marrow (BM) cells developed a polycythemia vera-like disorder. Treatment with the anti-oxidant N-acetylcysteine (NAC) substantially restored blood parameters and reduced damages to DNA. Furthermore, NAC induced a marked decrease in splenomegaly with reduction in the frequency of the Jak2(V617F)-positive hematopoietic progenitors in BM and spleen. Altogether, overproduction of ROS is a mediator of JAK2(V617F)-induced DNA damages that promote disease progression. Targeting ROS accumulation might prevent the development of JAK2(V617F) MPNs.