Industrialized seedling has the advantages of high efficiency, low cost, easy promotion of fine varieties, and standardized seedling management, so it is the trend of modern agriculture development. Seedlings using square substrate blocks is one of the main ways, but the high labor intensity and high labor costs have hindered its rapid development. In order to realize the automatic unloading of the seedling trays, this study designed and optimized a kind of seedling tray stacking manipulator for nursery machine that produces square substrate blocks. A design method for the robustness of grabbing the seedling trays and the anti-interference between manipulator and seedling tray was also proposed. Firstly, the principle of stacking manipulator was analyzed, and the structure was optimized. Kinematics analysis was conducted, and a kinematic model was established. The relationship between the stroke of an electric push rod and the opening and closing angle of a claw was obtained through model analysis. The movement stroke of the electric push rod was optimized to meet the design requirements of the maximum and minimum limit positions of the manipulator. Finally, the rationality of the parameter design was verified by static and kinematics simulation. The results demonstrate that the stroke range of the electric push rod is 27.6-105.6 mm, and the opening and closing angle range of the claw is 11.7-66.7°, which meets the requirements. In addition, the force of each component during the seedling moving is within a reasonable range. This study provides an important reference for the development and performance improvement of automatic square substrate seedling machines that support automatic loading, unloading, and palletizing.