Anoectochilus roxburghii is a perennial herb of the Orchidaceae family, used as a traditional medicinal material in China. Light is a major factor that affects the growth and quality of A. roxburghii. In this study, RNA-seq, GC-MS, and physicochemical analysis were employed to assess the developmental stages of A. roxburghii (S1: 30 d, S2: 60 d, S3: 90 d) under different light intensities (control light, CL: 1500 lx; strong light, SL: 4500 lx). There was no regular change in volatile oil content between developmental stages. Under strong light, 2-ethyl-5-methylfuranone content was significantly lower than that under control light. With the increase of cultivation age, the content of total flavonoids and polysaccharides exhibited an upward trend. Total flavonoid content under the strong light condition was significantly higher than that under control light. Transcriptome sequencing data yielded 86919 genes, which shared the highest homology with Dendrobium catenatum. In the S1 stage, there were 438 differentially expressed genes (DEGs) (SL/CL, upregulated: 190, downregulated: 248). These were mostly implicated in photosynthesis, chlorophyll metabolism, and sucrose metabolism. There were 468 DEGs in the S2 stage (SL/CL, upregulated: 378, downregulated: 90), mainly associated with sucrose metabolism, phenylpropane biosynthesis, and glucuronic acid transformation; 231 DEGs were observed in the S3 stage (SL/CL, upregulated: 73, downregulated: 158), implicated in plant-pathogen interaction, DNA replication, phenylpropane biosynthesis, and sucrose metabolism. Combining transcriptomic and physicochemical data suggested that light affects the expression of photosynthesis-related proteins, which in turn regulate glucose metabolism and flavonoid synthesis. This study provides a theoretical reference for changes in sugar metabolism and flavonoid biosynthesis in A. roxburghii under light stress, thus providing a basis for improving the quality of A. roxburghii.
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