Abstract: To construct Dendrobium officinale Kimura et Migo rapid propagation system，taking stems segments and leaves of Dendrobium officinale Kimura et Migo as explant，medium formulas of culture in vitro in different stages were designed.Results showed that optimum disinfection method was surface sterilization treatment of Dendrobium officinale Kimura et Migo with 70% ethanol 30s and with 0.1% Mercuric Chloride 9 min.Different explants on callus induction rate had big differences，the highest rate of stem segments of callus induction reached 83.3%.Optimum adventitious bud induction medium was MS+0.5 mgL 6-BA+0.2 mgL NAA；and rooting was 12 MS+0.5 mgL NAA.Optimum culture condition was determined，and a lot of callus and seedlings of Dendrobium officinale were gotten.It provides a scientific basis for establishment of rapid propagation system of Dendrobium officinale Kimura et Migo.