The invention provides a regulation method for a waterproof and drainage structure of a ballastless track subgrade. The regulation method comprises the following steps: chiseling away a first sealinglayer and a first grading macadam layer in a track-to-track area of a ballastless track; excavating a working well in the linear area, transversely jacking a drainage assembly into a base bed bottom layer of the subgrade, and filling seams in the periphery of the drainage assembly; arranging a water collecting well in the working well, and discharging liquid in the water collecting barrel out of the ballastless track, wherein the drainage assembly communicates with the water collecting well; pouring concrete in a mode of avoiding the base bed surface layer of the subgrade of the water collecting well in the track-to-track area to form a second sealing layer, wherein height of the second sealing layer is smaller than that of a supporting layer of the ballastless track; and sealing an embedded seam between the second sealing layer and the supporting layer. The regulation method solves the technical problem that drainage effect of an existing highly-sealed waterproof and drainage structure of the ballastless track subgrade is not good.