Establishment of an Efficient Rapid Propagation System and Transcriptomic Regulation Analysis of Curcuma kwangsiensis during Subculture and Transplantation

This study addresses the constraints hindering the development of the seedling industry, such as large seed usage and high production costs in rhizome propagation of Curcuma kwangsiensis , while research on in vitro rapid propagation of its plantlets remains scarce. Using C. kwangsiensis , a genuine medicinal material in Guangxi, as the experimental material, this study aimed to investigate in vitro regeneration and gene expression changes during culture via in vitro rapid propagation and RNA-Seq technology. Taking rhizome buds of C. kwangsiensis as explants, an efficient in vitro rapid propagation system was established by optimizing the media for cluster bud proliferation and rooting through single-factor tests and an L₉(3⁴) orthogonal experiment. Meanwhile, RNA-Seq was employed to analyze differential gene expression among the original plants, subcultured plantlets at different generations, and transplanted plantlets, so as to reveal the genetic variation patterns during tissue culture and transplantation. The results showed that the optimal medium for cluster bud proliferation of C. kwangsiensis was MS medium supplemented with 3.0 mg/L 6-BA, 0.9 mg/L TDZ and 0.7 mg/L NAA, with a proliferation coefficient of 5.13. The plantlets developed robust and well-proportioned root systems, and the transplantation survival rate reached 98%. Plant height, leaf length, proliferation coefficient and root number of the tissue-cultured plantlets decreased significantly with the increase of subculture generations, and obvious degeneration occurred after the 6th subculture (T6). Transcriptomic analysis indicated that the number of differentially expressed genes (DEGs) increased with the rise of subculture times, and these DEGs were mainly enriched in pathways such as cellular process, metabolic process, plant hormone signal transduction and phenylpropanoid biosynthesis. The gene expression pattern returned to that of the original plants after transplantation. This study established an efficient in vitro rapid propagation system for C. kwangsiensis , clarified the phenotypic and molecular mechanisms underlying subculture-induced degeneration and transplantation-mediated recovery, and provided a theoretical basis and technical support for high-quality seedling breeding, as well as germplasm purification and rejuvenation of C. kwangsiensis .