The invention discloses a high-frequency differentiation medium of Caoyulu callus, which is composed of inorganic salt, organic components, carbon source, additives, coagulant and hormone, wherein the inorganic salt contains NH 4 NO 3 , KNO 3 , CaCl 2 , KH 2 PO 4 , MgSO 4 , KI, etc., the organic components contain Na 2 EDTA, inositol, niacin, thiamine hydrochloride, pyridoxine hydrochloride and glycine, the carbon source is sucrose, the additive is Caoyulu callus extract, the coagulant is crystal agar, and the hormones are 6‑BA and NAA. The medium can significantly improve the differentiation ability of cells, realize the high-frequency differentiation and regeneration of callus, and form a well-grown grass-gyulu plant under the condition of no root system. There is no need to change the medium type during the entire culture period, which significantly shortens the The time for callus redifferentiation is shortened, and a large number of easy-to-survive tissue culture seedlings are obtained, and the cultivation process is not limited by seasons, which effectively improves the reproduction efficiency and economic benefits.