Nitrous oxide (N2O) is a potent greenhouse gas and of special concern in wastewater treatment. It is formed in biological wastewater treatment under both aerobic and anaerobic conditions. A major reason for high N2O emissions is low oxygen concentration during nitrification. In this full-scale study of N2O emissions from a sequencing batch reactor for treating digester supernatant, the oxygen concentration was reduced stepwise to investigate how N2O emissions were influenced. N2O concentrations were measured online in water and off-gas. A distinct relationship was found between low oxygen concentration and high N2O emissions. N2O was formed in water during both nitrification and denitrification. Decreased oxygen concentration during nitrification led to increased nitrite concentration, which in turn led to increased N2O concentration in the subsequent denitrification phase. When the nitrification resumed, accumulated N2O was stripped off to the atmosphere. Very high concentrations of N2O, over 56,000 ppmv, were measured in the off-gas. Furthermore, the maximum amount of N2O emitted during one cycle corresponded to 107.6% of the total nitrogen load (21.9% of total nitrogen present in the bulk liquid at the beginning of the cycle). This is among the highest emission levels ever measured from a full-scale municipal plant for digester supernatant.