In this work, an ultrasensitive and highly stable non-enzymatic glucose sensor is developed with CuO nanostructures through a facile, low price, wet-chemistry route. Three kinds of CuO nanostructures are obtained by kinetic control. The best as-prepared CuO electrode with rough petal-like structure exhibits a quantitative sensitivity as high as 7546.37 μA cm−2 mM−1 and an ultra low limit detection 0.259 μM, which are much better than some previously reported. The rough petals might augment the contact area for kinetic mass transfer between CuO nanostructures and the electrode in an alkaline environment, which significantly improves the performances of electro-oxidation of glucose. Furthermore, the as-fabricated non-enzymatic glucose sensors have also shown excellent anti-interference property, fast response time, good reproducibility and long duration stability. Thus, as-fabricated CuO electrode can be a promising material in the application of non-enzymatic glucose sensors. Display omitted •An ultrasensitive non-enzymatic glucose sensors based on controlled petal-like CuO nanostructure were obtained.•The best as-prepared CuO electrode (Nafion/CuO8/GCE) showed an excellent sensitivity of 2634.44 μA cm−2 mM−1 in within a wide linear range from 0.5 μM to 2.67 mM.•The Nafion/CuO8/GCE electrode also shows excellent limit of detection, fast response time, good reproducibility and long duration stability.•The outstanding properties of Nafion/CuO8/GCE electrode may ascribe to (i) the morphologies (ii) strong binding of Tween 80 (iii) rough surface.