Stage and Cultivar Specific Regulation of Cannabinoid in Hemp: An Integrated Omics Perspective

we employed integrated transcriptomic and metabolomic analyses to elucidate the molecular basis of cannabinoid biosynthesis during early and late flowering stages in two hemp cultivars, CBG-Ambit and CBG-Limonene. A total of 338 differentially expressed genes (DEGs) were identified at the early stage (ES), while 342 at the late stage (LS) between CBG-Ambit (Amb) and CBG-Limonene (Lim) cultivars. Additionally, 1342 DEGs were identified between the early and late stages of Ambit, whereas 1152 DEGs among early and late stages in Limonene. Key candidate genes including cytochrome P450 78A9 (CYP78A9), 3-hydroxy-3-methylglutaryl-coenzyme A reductase 1 (HMGCR1), auxin transporter-like protein 2 (LAX2), myrcene synthase (MyrS), terpene synthase 29 (TS-29), geranylpyrophosphate: olivetolate geranyltransferase (GPP), ABSCISIC ACID-INSENSITIVE 5-like protein 7 isoform X1 (ABI5), MYB transcription factor 1, histone-lysine N-methyltransferase (SUVH6), TIFY 10A, tetraketide alpha-pyrone reductase 2 (TPKR2), ethylene-responsive transcription factor (ERF109), berberine bridge enzyme-like 8 (BBE8) and monoterpene synthase (MTS1), were implicated as central regulators of CBD biosynthesis and conformed by RT-qPCR. Functional enrichment (GO and KEGG) analysis demonstrated that these genes are concentrated in secondary metabolic processes, oxidoreductase activity, and transcriptional regulation, highlighting their central roles in cannabinoid and terpenoid biosynthesis. Along with ultra-performance liquid chromatography-mass spectrometry (UPLC-MS), metabolomic profiling further showed increased accumulation of CBDA and CBGA in late stage. Identification of differentially expressed genes in early and late stage of CBG-Ambit and CBG-Lim for cannabinoid biosynthesis