Abstract: Transplanting machines have been widely used for the chrysanthemum seedlings in smart agriculture. Challenges have remained on the unstable seedling gripping damage to seedlings. Fortunately, a three-needle concentrating seedling picker can be expected for the seedling extraction. However, current models often focus primarily on the growing medium rather than the root fixation. This study aims to construct the three-dimensional complex root system of chrysanthemum seedlings using L-system theory. A numerical model was established to more accurately simulate the root architecture and its interaction with the surrounding medium during extraction. A EDEM-RecurDyn model was coupled with discrete element method and multi-body dynamics. The mechanical behavior of the seedlings was predicted to experimentally validate interaction between the complex root system, the growing medium, and the seeding needles during seedling extraction. A systematic investigation was made to explore the impact of various operational factors, such as the structure of seeding needles, seeding depth, seeding speed, and push plate stroke on the mechanical properties and damage levels of the seedlings. The seeding needle and operational parameters were finally optimized to minimize the seedling damage with the high success rates during seedling extraction. The results indicate that the seedling damage depended on the seeding depth, speed, and push plate stroke. Notably, the seeding depth increased the seedling displacement, which in turn increased the risk of damage, while the higher seeding speed caused more severe breakage of root system. The push plate stroke also dominated the movement of the seedling within the claw, the overall stability and force distribution during extraction. Multivariable regression was developed to predict the relationship between operational variables—seeding depth, seeding speed, and push plate stroke—and the seedling’s damage rate and integrity using Response Surface method. The optimal extraction parameters were determined to be an seeding depth of 20 mm, an seeding speed of 200 mm/s, and a push plate stroke of 5 mm. The extraction success rate reached 84.3%, with the seedling integrity rate of 93.10%. The optimal parameters were significantly reduced the seedling damage during extraction. Field trials were conducted to further validate the optimization for the high seedling quality. The operational settings were verified for the transplanting success rates after simulation. The optimal seeding needle was reduced seedling damage for the overall performance of the transplanting machine. In conclusion, the seedling extraction mechanisms were developed to consider the mechanical interactions among seeding needles, seedling root system, and the growing medium during extraction. Three-pronged seeding needle was coupled with the optimal parameters to highlight the seedling fixation, including root characteristics. The finding can offer the strong reference for the high transplanting success rates and seedling quality in transplanter.