Abstract: In this study, in vitro culture system of Dracocephalum rupestre Hance was established for the first time by evaluating the effects of different explants, different types and concentrations of plant hormones on bud induction,proliferation and adventitious root induction. The results indicated that the stem with axillary buds was the most suitable explants for Dracocephalum rupestre Hance tissue culture and rapid propagation; the optimum medium for bud induction was MS ＋6-BA 0.5 mg/L ＋NAA 0.1 mg/L ＋IAA 0.2 mg/L, and the average induction rate was 63.6%; the proliferation culture medium for bud differentiation was MS＋6-BA 0.75 mg/L＋KT 4.0 mg/L＋NAA 0.01 mg/L＋ GA3 0.2 mg/L, with the average single bud proliferation number being 4.3; the optimum adventitious root induction medium was MS＋NAA 0.2 mg/L, and the rooting rate was 85.1%; the transplanting survival rate reached 100%. The results laid the foundation for the industrial production of Dracocephalum rupestre Hance plantlets.