Aiming at the problems that the roots and stems of mechanized transplanting of bare rape seedlings are intertwined, which makes it difficult to separate seedlings and the conventional seedling separation device is easy to damage the roots and stems, a seedling separation process of artificial assisted row placement + flexible clamping transportation + intermittent dial separation was proposed, and a flexible clamping seedling separation device was developed. A flexible clamping seedling separation device was designed. The material characteristics of rapeseed bare seedlings during the optimal transplanting period were measured, determining the transverse conveyor belt width of the flexible clamping mechanism to be 216 mm. The dynamics of rapeseed bare seedlings during the flexible clamping and conveying process were analyzed, and the feed inlet angle was determined to be 30°. The trajectory of manual seedling separation was simulated, and rigid-body guidance of the separation mechanism was analyzed based on Burmester theory, resulting in the determination of a crank length of 32 mm and a rocker length of 50 mm. Motion parameter matching analysis was conducted for the flexible clamping mechanism and the seedling separation mechanism, determining the flexible clamping mechanism's conveying speed to be 25~55 mm/s and the separation mechanism's poking frequency to be 50~120 times/min. An orthogonal bench test was conducted with the separation outlet angle, the conveying speed of the flexible clamping mechanism, and the poking frequency of the separation mechanism as test factors, and the seedling separation success rate and seedling damage rate as evaluation indices. The test results showed that the optimal parameter combination was a separation outlet angle of 55°, a conveying speed of 48 mm/s for the flexible clamping mechanism, and a poking frequency of 85 times/min. Under these conditions, the seedling separation success rate and damage rate of the device were 89.06% and 3.91%. The field test showed that each mechanism of the seedling separation device ran smoothly, and the operation performance met the requirements of mechanized transplanting of bare rape seedlings. The research results can provide technical reference for the design of automatic transplanting equipment for bare rape seedlings.