The interfacial assembly of high roughness thin-layer noble metal nanostructures is important for the development of high performance electrochemical sensors. Herein, by electrodepositing a semiconductor Te film with sawtoothed shape on the glassy carbon electrode (GCE) as the template and a subsequent galvanic replacement reaction with HAuCl4, the bar-shaped and rough Au thin film modified glassy carbon electrode (AuTe-R/GCE) was fabricated. The electrocatalytic performance of the modified electrode toward glucose oxidation was studied in alkaline condition via cyclic voltammetry, and the glucose non-enzymatic amperometric sensor was constructed. Compared with the conventional gold-electroplated GCE (Aucon/GCE) and bare Au electrode, the AuTe-R/GCE showed the higher electrocatalytic activity toward the oxidation of glucose. Under the optimal conditions, the glucose concentration was detected via potentiostatic chronoamperometry, and the AuTe-R/GCE showed a linear detection range of glucose concentration from 0.01 to 2.00 mmol/L with a sensitivity of 3.8 mA mmol-1 cm-2, and limit of detection (LOD) of 55 nmol L-1, with strong anti-interference ability and good stability, being superior to most of relevant reports. The new method for preparing the specific morphology and high roughness thin-layer nano-sized Au by replacement with semiconductor Te film presented here has the advantages of convenience, rapidness and low-cost, and is expected to be widely used in the interfacial assembly of highly active nanoelectrocatalysts and their electrochemical performances research.