Glycogen derivative with strong positive was synthesized by two steps. Firstly, the glycogen was modified by ethylenediamine to obtain the aminated glycogen derivative (N-Gly). Secondly, N-Gly was modified by 2, 3-epoxypropyl trimethyl ammonium chloride (GTA) to obtain the cationic quaternary ammonium glycogen derivative (QA-N-Gly). Then, quaternary ammonium glycogen derivative-gelatin composite hydrogel (QA-N-Gly/Gel) was prepared by amide central crosslinking. The chemical structure of glycogen derivative was characterized by FTIR and 1H-NMR. The particle size distribution, Zeta potential and morphology of glycogen derivative were also investigated. The results showed that glycogen was successfully aminated and quaternary ammonium modified. The QA-N-Gly were dispersed nanoparticles with a size distribution of 100-200 nm and a Zeta potential of 90-100 mV. QA-N-Gly exhibited antibacterial effect because of significant positive charge. The physicochemical and biological properties of QA-N-Gly/Gel hydrogel were evaluated. The results showed that the addition of glycogen derivatives could improve the mechanical strength and stability of gelatin-based hydrogel. The fracture stress of QA-N-Gly/Gel was 66.8 KPa, and the compression modulus was 13.3 KPa. QA-N-Gly/Gel showed obvious antibacterial effect on Staphylococcus aureus and Escherichia coli, and the inhibition rate reached 97% and 50%, respectively. In addition, QA-N-Gly/Gel showed no hemolysis and low cytotoxicity, the 48 h survival rate of NIH-3T3 cells was more than 80%.