Abstract: Microbial exopolysaccharide (EPS) is an important biological resource, and EPS production from endophytes has attracted much attention in recent years. To explore more EPS-producing strain resources as well as soybean endophytic resources, in this study, the strain with the highest EPS yield was screened from soybean seeds, and its morphological and molecular identification, biological characteristics (temperature, pH, rotation speed, sucrose tolerance, NaCl tolerance) and physiological and biochemical studies were carried out. The results showed that the maximum EPS yield of strain SE01 was 0.63 g/L after 48 h fermentation. Colony of strain SE01 was round, with smooth edge, milky white, and glossy and sticky surface. Under microscope, the bacterium after Gram staining was short, pink and rod-shaped, which identified to Gram-negative bacterium. The results of physiological and biochemical experiments and 16S rDNA identification showed that strain SE01 was Enterobacter cloacae, and named Enterobacter cloacae SE01. The optimum growth conditions of Enterobacter cloacae SE01 were temperature 30 ℃, pH5, and rotational speed 120 r/min. The strain SE01 had high tolerance to sucrose, but low tolerance to NaCl. Increasing the concentration of NaCl could inhibit its growth. This study provides data support for the development and application of soybean endophytic EPS.