Daphnia normally reproduce by cyclical parthenogenesis, with offspring sex being determined by environmental cues. However, some females have lost the ability to produce males even in the face of strong hormonal stimuli. Our results demonstrate that this loss of male-producing ability is subject to genetic control by a dominant allele at a single locus. We identified the non-male-producing locus by comparing whole-genome sequences of 67 non-male-producing (NMP) and 100 male-producing (MP) clones from five Daphnia pulex populations. We identified 132 NMP linked markers within a single 1.0 Mb nonrecombining region on chromosome I. These markers include seven nonsynonymous mutations, all of which are located within one unannotated protein-coding gene .