Abstract: To explore the suitable cultivation substrate for the growth of test-tube seedlings of Cymbidium spp. ‘Ranluo’. Tissue cultured seedlings of ¹Cymbidium spp. ‘Ranluo’ were used as materials to analyze their effects on survival rate, plant height, pseudobulb size, root length, root volume, etc. by different substrate combinations including pine bark, Cedar bark, bamboo shavings, peat soil, perlite, etc. The results showed that in terms of survival rate, except for pine bark and bamboo shavings, the survival rate of other combination treatments could reach 100%. In terms of plant height, false bulb size, and root length, the Cedar bark: peat soil=4∶1 treatment performed the best, with significant differences compared to other groups. In terms of root volume, the Cedar bark: perlite=4∶1 and Cedar bark: peat soil=4∶1 treatments performed the best. Based on the comprehensive survival rate, plant height, pseudobulb size, root length, and root volume, a ratio of 4∶1 between Cedar bark and peat soil is more conducive to the growth of test tube seedlings of the Cymbidium spp. ‘Ranluo’, providing guidance for the production practice of the Cymbidium spp. ‘Ranluo’.