Chelating agent is widely used in industries such as papermaking, dyeing and food etc.. The instability and toxicity of it are harmful to environment and human beings. It is important and significant to find an environmentally friendly chelating agent. Strain WJ-3 was used to synthesize siderophore. The effects of carbon source, nitrogen source, temperature, shaking speed, and pH on the initial medium and siderophore were studied by single factor method. The results showed that strain WJ-3 could synthesize siderophore both in solid and liquid medium. The optimal carbon and nitrogen source were glucose and yeast extract powder. The optimal cultivation conditions were with temperature at 30 ℃, 180 r/min for shaking speed, and pH 7.0 in the initial medium. The synthetic rate of microbial siderophore could reach 64.12% after shaking culture for 48 h. It suggested that the optimal conditions could increase the synthesis of microbial siderophore.