Establishment of an In Vitro Regeneration System and Analysis of Endogenous Hormone Dynamics in Melastoma dodecandrum

Melastoma dodecandrum is primarily propagated through stem cuttings, which limits genetic variation and constrains breeding efforts. To overcome this limitation and facilitate molecular breeding, the establishment of a reliable and efficient regeneration system is essential. This study investigated the effects of plant growth regulators (PGRs) and culture media on the in vitro regeneration system of M. dodecandrum. The highest rate of callus induction (96.67%) was achieved when sterile leaf explants were cultured on Murashige and Skoog (MS) basal medium supplemented with 2.00 mg&middot:L&minus:1 2,4-dichlorophenoxyacetic acid (2,4-D) and 0.50 mg&middot:L&minus:1 6-benzylaminopurine (6-BA). For callus differentiation, the optimal formulation of MS + 2.0 mg&middot:L&minus:1 6-BA + 0.5 mg&middot:L&minus:1 naphthylacetic acid (NAA) resulted in a differentiation frequency of 83.33%. The optimal PGR combinations for shoot proliferation were 1.5 mg&middot:L&minus:1 6-BA + 0.1 mg&middot:L&minus:1 NAA and 0.5 mg&middot:L&minus:1 6-BA + 0.2 mg&middot:L&minus:1 NAA. The optimal rooting media were MS medium supplemented with 0.1, 0.2, or 0.5 mg&middot:L&minus:1 indole-3-butyric acid (IBA) or 1/2MS medium supplemented with 0.1 mg&middot:L&minus:1 IBA. Additionally, this study investigated the dynamic changes in endogenous hormones during the regeneration process. The levels and ratios of hormones, including gibberellin (GA3), abscisic acid (ABA), indole-3-acetic acid (IAA), and zeatin (ZT), collectively regulated the regeneration process. Elevated levels of ABA and GA3 may promote callus initiation as well as the growth and development of adventitious roots during the early induction stage. Reduced levels of ABA and IAA favored callus differentiation into shoots, whereas elevated GA3 levels facilitated proliferation of adventitious shoots. Throughout the regeneration process, fluctuations in ZT levels remained relatively stable. This study successfully established an in vitro regeneration system for M. dodecandrum using leaf explants, providing theoretical guidance and technical support for further molecular breeding efforts, genetic transformation, and industrial development.