Abstract: In order to select the suitable culture substrate for the domestication and transplanting of Dendrobium officinale tissue cultured seedlings, 9 different substrate combinations were used to carry out the transplanting test, in order to screen out the suitable culture substrate. The results showed that the effects of different matrix combinations on the survival rate, incidence and growth of tissue cultured seedlings of Dendrobium officinale were significantly different. The survival rate of treatment 5 (crushed stone∶mosses= 1∶2) was the highest, which was significantly higher than that of other treatment groups, and the incidence was the lowest, which was significantly lower than that of other treatment. The survival rate and incidence were 96.92% and 1.1%, respectively. In addition, the six growth indexes of Dendrobium officinale in the treatment 5, including the fresh weight, plant height, stem thick, budding number, total root number, and the longest root length were also significantly higher than those in the other treatment, which respectively increased by 40.74%, 48.43%, 7.69%, 5.86%, 11.70% and 3.97% compared with the control (fine shaving wood chips 5 + sawdust 1). It was the best substrate for the domestication and transplanting of tissue cultured seedlings of Dendrobium officinale.