## 2 Results
### Functional Analysis of DEGs
To identify genes closely associated with terpenoid biosynthesis under light and darkness treatments, we screened DEGs based on the L2 vs. D2 comparison. Using the WGCNA method in the R package (R version 4.2.1), including FPKM values from L1-L4 and D1-D4, we constructed a weighted gene co-expression network with a cutoff value >0.5 and visualized the network using Cytoscape (v3.8.0). In Figure 6C, we identified the _Lcu01G_23775_ gene as a key ADH (Alcohol Dehydrogenase) terpene synthase gene, which exhibited extremely high correlations with four transcription factors. ADH functions to catalyze the formation of terpene skeletons and subsequent oxidation/reduction modifications, the final step in citral biosynthesis in _L. cubeba_. The four transcription factors belong to the NAC (_novel.2618_ and _novel.4226_) and WRKY (_Lcu08G_23653_ and _Lcu08G_23775_) gene families, which are widely involved in regulating plant secondary metabolic processes (Figure 6C) [49; 50; 51]. Figure 6: GO, KEGG, and WGCNA analysis of the DEGs (L2 vs. D2). (**A**) GO enrichment analysis of the DEGs (L2 vs. D2) was categorized into biological process, molecular function, and cellular component. (**B**) The DEGs (L2 vs. D2) were enriched in five KEGG metabolic pathway categories. Different colors represent different categories, and the numbers after the pathways represent the number of single genes in the different categories. (**C**) Construction of regulatory networks of TFs and structure genes related to terpenoid biosynthesis. Yellow represents the key enzyme ADH in the terpenoid synthesis pathway, and purple represents transcription factors. The Expression Profile of Genes in the Terpenoid Biosynthetic Pathway in Under Light and Darkness Treatments  
To elucidate the changes in the expression of key genes in the terpenoid synthetic pathway in _L. cubeba_ under light and darkness treatments, we constructed a schematic diagram of this pathway, incorporating the transcriptomic profiles of critical enzymatic nodes (Figure 7). Within the MVA pathway, key biosynthetic genes (_LcACOT2_, _LcACOT3_, _LcPMK_, _LcID11_, _LcTPS5_, _LcTPS14_) exhibited marked transcriptional upregulation under photoperiod conditions relative to darkness-treated controls. This light-responsive regulatory pattern was similarly observed in the MEP pathway, where _LcDXS1_, _LcDXS3_, _LcHDR2_, _LcTPS21_, _LcTPS23_, _LcTPS27_, _LcTPS28_, _LcTPS29_, _LcTPS30_, _LcTPS31_, _LcTPS32_, _LcTPS33_, and _LcTPS34_ demonstrated significant upregulation under light treatment compared to darkness treatment. Conversely, genes such as _LcDXR_, _LcMDS_, _LcHDR1_, _LcGPPS2_, _LcTPS18_, _LcTPS19_, and _LcTPS20_ exhibited elevated expression levels in the darkness treatment process. In addition, we found that under the darkness treatment (D3-D4), the expression levels of most genes in the MVA and MEP pathways, including those in the _HMGS_, _MK_, _PMK_, _FPPS_, _DXS_, _DXR_, and _HDS_ gene families, were significantly decreased. Notably, the expression levels of _LcACOT1_, _LcHMSG1_, _LcHMR3_, _LcDNC_, _LcDXS1_, _LcDXS6_, _LcCMS_, _LcHDS1_, _LcHDS2_, and _LcGPPS2_ exhibited strong positive correlations with monoterpene accumulation dynamics during darkness treatment (Figure 7). These findings indicates that transcriptome sequencing represents an important method for screening candidate genes that can enhance terpenoid production in breeding programs. Figure 7: Schematic diagram of the terpenoid biosynthetic pathway in _L. cubeba_ under light and darkness treatments (L1–L4 and D1–D4).