One of the important stages of plants propagated by tissue culture before being moved to the field is acclimatization. Small plants must adapt to the outside environment in the acclimatization room. This research was aimed to figure out the composition of growing medium with the best concentration of auxin for the growth of chrysanthemum plant in the acclimatization room. This research was conducted from February until April 2017 in Hargobinangun Village, Pakem District, Sleman Regency, Special Region of Yogyakarta Province. The research was a field experiment with factorial Completely Randomized Design and was repeated five times. The growing medium tested was vermin compost (kascing fertilizer), manure and compost. Meanwhile, the concentrations of auxin being tested were 1.2 and 3 ppm. The results of the research show that the growing medium of compost made from bamboo leaves could increase the number of leaves, the height of the plant, the number of roots and the fresh weight of the plant. Meanwhile, the concentration of auxin of 2 ppm was able to increase the height of the plant, the number of roots, the length of roots, and the fresh weight of the plant.