Cnidoscolus aconitifolius (Mill.) I.M. Johnst is a multipurpose woody plant. In this study, an in vitro efficient propagation system of stem segment explants derived from field-grown C. aconitifolius plants was established for the first time. The sterilization effect, axillary bud initiation, and proliferation efficiency of stem segments were evaluated. The results showed that the sterilization time of 0.1% mercuric chloride, the concentration of Plant Preservative Mixture (PPM), the pretreatment method, and the sampling season had significant effects on the sterilization of stem segments (p < 0.05). The type of medium and plant growth regulators (PGRs) affected the initiation of axillary buds, and the proliferation efficiency was significantly affected by PGRs. The results showed that the best sterilization method for stem segment explants was as follows: a pretreatment by rinsing with running water for 120 min, soaking in 75% ethanol for 50 s, soaking in 0.1% mercuric chloride for 10 min, and medium supplemented with 3 mL/L PPM. When inoculated on the medium in spring, the contamination rate was as low as 25.56%. The optimal initiation medium for axillary buds in stem segments was half-strength Murashige and Skoog (1/2 MS) medium supplemented with 0.5 mg/L 6-benzyladenine (6-BA). The induction rate was as high as 93.33%, and the mean length of axillary buds was 2.47 cm. The optimal proliferation medium was 1/2 MS medium supplemented with 4.0 mg/L 6-BA and 0.2 mg/L indole-3-butyric acid (IBA). The induction rate was up to 80.00%, the total proliferation coefficient was 4.56, and the net proliferation coefficient was 5.69. The 1/2 MS medium supplemented with 0.1 mg/L 6-BA and 1.5 mg/L indole-3-acetic acid (IAA) was most conducive to the elongation of the adventitious shoot, and the adventitious shoot of approximately 1 cm reached 1.93 cm after culturing for 14 days. The best medium for adventitious shoot rooting was 1/2 MS medium supplemented with 0.1 mg/L α-naphthalene acetic acid (NAA), the highest rooting rate was 82.00%, and the survival rate of transplanting was over 90%.
Keywords: Cnidoscolus aconitifolius; aseptic system; plant growth regulators; tissue culture.