Mouse oocyte meiotic division requires the establishment of asymmetries in the oocyte before division, indicating the presence of polarity-establishing molecules. During mouse oocyte maturation proper orientation and positioning of the meiotic spindle at the oocyte cortex, as well as polarity in the oocyte cytoplasm and its oolemma, are necessary for the formation of functional haploid oocytes. Discs large homologue 1 (Dlg1) protein is a conserved protein that regulates cell polarity. In the present study, we found that Dlg1 was expressed at different stages of oocyte development. The localisation of Dlg1 during mouse oocyte maturation and its relationship with the cytoskeleton were analysed. Our data show that at the germinal vesicle stage, Dlg1 was present in the cytoplasm, prominently surrounding the germinal vesicle membrane. During maturation, Dlg1 became highly polarised by associating with the spindle and formed characteristic crescent-shaped accumulations under the cortex. Addition of nocodazole or cytochalasin B into the culture medium at different stages changed the localisation of Dlg1, indicating that the organisation of Dlg1 is a complex multi-step process and is dependent on microtubules and microfilaments. More importantly, we found that silencing of Dlg1 compromised the G2-M transition.