Haplotype-resolved cannabis genome reveals hidden allelic drivers of medicinal compound diversity

A fully phased, chromosome-level genome assembly of a wild Tibetan cannabis accession has revealed that extensive genetic differences between the two inherited chromosome copies—known as haplotypes—are a major driver of the plant's diverse medicinal compound profile. Researchers found that structural variations and transposable element insertions differentially shape the architecture of key biosynthetic genes, leading to divergent expression and function between haplotypes. This allelic divergence directly influences the production of cannabinoids, terpenoids, flavonoids, and other specialized metabolites, offering a new framework for understanding how genetic diversity within a single plant generates its therapeutic and ecological chemistry.

Cannabis sativa produces hundreds of specialized metabolites, including cannabinoids, terpenoids, and flavonoids, many of which hold significant therapeutic promise. However, the plant's highly heterozygous genome—meaning the two sets of chromosomes inherited from each parent are substantially different—has long obscured a complete understanding of how these compounds are biosynthesized and regulated. Traditional genomic studies that collapse both haplotypes into a single reference sequence fail to capture the distinct contributions of each chromosome copy, masking the allele-specific variations that are fundamental to gene expression and trait inheritance. Based on these challenges, a haplotype-resolved approach that distinguishes between the two genomes within a single plant is needed to dissect the regulatory complexity underlying cannabis metabolism.

A team led by researchers at Fujian Agriculture and Forestry University, in collaboration with Yunnan University and Israel's Volcani Institute, has generated a fully phased, chromosome-level genome assembly of a wild cannabis accession from Tibet. Publishing (DOI: 10.1093/hr/uhag159) in 2026 in Horticulture Research, the study integrates comparative genomics, transcriptomics, and structural variant analysis to reveal how haplotype-specific variation drives the biosynthesis of cannabinoids, terpenoids, flavonoids, and other specialized metabolites across different tissues and developmental stages.

The researchers assembled two complete haplotype genomes, HapA and HapB, totaling 1.48 gigabases, and identified extensive structural asymmetry between them—including more than 32,000 insertions, nearly 117,000 duplicated sequences, and thousands of inversions and translocations. Many of these structural variants directly impact genes encoding key metabolic enzymes. At the germacrene D synthase (GERD) locus, for example, the HapB allele carries both a truncation that likely abolishes enzyme function and a promoter mutation disrupting a MYC transcription factor binding motif, rendering it transcriptionally silent while the HapA allele remains active. Similarly, the oxidosqualene cyclase (OSC) gene family exhibits haplotype-specific copy number variation—three active copies in HapA versus one silent copy in HapB—with functional validation in Nicotiana benthamiana confirming β-amyrin production from the active HapA copy. Transposable element (TE) insertions were found to exert a pervasive suppressive effect on gene expression, with TE-associated genes showing significantly lower expression across all tissues. Notably, a long terminal repeat (LTR) insertion in the intron of a lipid transfer protein gene (LTP4-like) in HapB correlates with transcriptional repression, while three consecutive TE insertions in the intron of olivetol synthase (OLS) in HapA likely drive near-complete silencing of that allele, directly shaping cannabinoid precursor production in glandular trichomes.

"The extent of functional divergence between the two haplotypes really surprised us," the authors said. "We're not just seeing different expression levels—we're seeing alleles that have completely lost enzymatic activity, others that have been silenced by transposon insertions, and gene families that have expanded on one haplotype while remaining static on the other. It suggests that wild cannabis maintains this allelic diversity as a strategy, potentially allowing the plant to draw from a broader biochemical toolkit to adapt to changing environments. This is a level of regulatory complexity that you simply cannot see without a phased genome."

This haplotype-resolved framework provides a foundation for genomics-guided breeding and metabolic engineering in cannabis. By identifying which specific alleles drive the production of high-value cannabinoids and terpenes, breeders can selectively favor favorable haplotypes to optimize medicinal compound profiles without compromising other agronomic traits. The findings also highlight the importance of preserving genetic diversity in wild germplasm collections, as wild accessions like CSLZ harbor unique allelic variants that have been lost or diminished in domesticated varieties. Beyond cannabis, this study demonstrates how haplotype-aware genomic approaches can uncover hidden layers of regulatory variation underlying metabolic diversity in other highly heterozygous crops and medicinal plants.

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References

DOI

10.1093/hr/uhag159

Original Source URL

https://doi.org/10.1093/hr/uhag159

Funding information

National Natural Science Foundation of China (32561143033, 32470281); Natural Science Foundation for Distinguished Young Scholars of Fujian Province (2025J010027) [40†L868-L869].

About Horticulture Research

Horticulture Research is an open access journal of Nanjing Agricultural University and ranked number one in the Horticulture category of the Journal Citation Reports ™ from Clarivate, 2023. The journal is committed to publishing original research articles, reviews, perspectives, comments, correspondence articles and letters to the editor related to all major horticultural plants and disciplines, including biotechnology, breeding, cellular and molecular biology, evolution, genetics, inter-species interactions, physiology, and the origination and domestication of crops.