Chris Grassa Thesis Defense (James Mallet, Advisor)

Title: Establishing a Genetic System for Studying Cannabis Domestication

Abstract: Cannabis is a globally significant crop with a long history of domestication for fiber, food, and psychoactive compounds. Despite its economic and cultural importance, a comprehensive understanding of its evolutionary history, the genetic basis of its diversification, and the selective pressures that shaped its genome remains incomplete. In my dissertation, I employed an interdisciplinary approach integrating genomics, population genetics, molecular dating, and cultural-economic analysis to resolve key questions in the evolution and domestication of Cannabis. In Chapter 1, I established the theoretical and historical context for this work with a comprehensive literature review. This chapter synthesizes the multi-stage process of Cannabis domestication, traces its historical geography and diversification across Eurasia, and details the biology of key agronomic traits under selection, including seed dehiscence, sexual expression, flowering time phenology, and trichome density. In Chapter 2, I generated and analyzed a chromosome-scale reference genome for a high-cannabidiol (CBD) cultivar. Quantitative trait locus (QTL) mapping in a hemp × marijuana cross identified a single locus on chromosome 7 that explains >90% of the variance in the THC:CBD ratio. Population genomic analysis revealed that the high-CBD phenotype results from the recent introgression of a functional hemp-derived CBDAS allele into a high-potency marijuana genetic background, which typically possesses a non-functional THCAS allele, demonstrating rapid evolution in response to changing legal frameworks. In Chapter 3, I constructed a fossil-calibrated molecular chronogram for the Cannabaceae family using a comprehensive phylogenomic dataset. Through rigorous re-evaluation of fossil evidence, this analysis established a robust temporal framework for the family's evolution, estimating the divergence of Cannabis and its sister genus Humulus at 35.6 (95% HPD: 33.94–38.30) million years ago, providing critical context for the deep-time evolution of the lineage. In Chapter 4, a population genomic analysis of wild and feral Eurasian Cannabis identified ten major polymorphic chromosomal inversions. These structural variants exhibit strong clinal variation across latitudinal gradients, contain genes enriched for functions related to flowering time and environmental stress response, and show significant inter-inversion linkage disequilibrium. These results indicate the inversions function as supergenes that facilitate local adaptation and maintain ecological differentiation between populations. Finally, in Chapter 5, I quantified the selective pressures of 20th-century prohibition on modern drug-type Cannabis. Economic analysis revealed a strong positive correlation (R² ≈ 0.9) between federal law enforcement expenditure and THC potency, an adaptive market response consistent with the Alchian-Allen Theorem. This chapter also documents extreme artificial selection on the Cannabis sexual system, where the estimated selection coefficient (s) against males approaches 1.0 in elite cultivation, resulting in functionally unisexual populations maintained via clonal propagation and feminized seed. Collectively, this dissertation provides a multi-scale analysis of Cannabis evolution. It establishes a refined evolutionary timeline, identifies large-scale structural variants as key drivers of adaptation, and quantifies the intense, recent selective pressures that have produced the modern crop

Committee: James Mallet (Advisor), David Haig, Elena Kramer