## 1 Introduction
Aside from CBs, the cannabis plants also produce other specialized metabolites from shared precursors with cannabinoids, such as fatty acids, competing with olivetolic acid synthesis, and terpenoids such as limonene [23], competing for the GPP pool [24; 25]. The yield of CB in plants is limited, both in quality and in quantity; the maximum production is due to a mixture of these metabolites accumulating in the flowers (20 mg/g fresh weight) [6; 26]. In addition, the lack of scientific literature on cannabis physiology leads to the absence of standardized culturing protocols, resulting in a variation in CB profiles, quality, and quantity. In addition, cannabis pathogens such as the gray mold causal agent (_Botrytis cinerea_) affect the overall plant yield and metabolite composition [27]. As well as multiple unknown facets of CB production in planta, this suggests that the implementation of a heterologous expression system with fewer components and fewer competing enzymes might be a solution for achieving a higher production yield of single CB components, such as CBD or THC. Therefore, synthetic biology offers an important alternative method to produce CBs for the pharmaceutical field with less risk and fewer crop management challenges. Several research groups aimed to design de novo or improve pre-existing cannabinoid production in plant tissue cultures, but apart from the transformation rates, in vitro culture viability and yield are still to be optimized [6]. For instance, few groups succeeded in transforming yeast (_Saccharomyces cerevisiae_) and bacteria (_Escherichia coli_) to produce the different metabolites of the CB pathway, such as OA or CBGA [28; 29]. The production of these metabolites in yeast and bacteria often involves the addition of costly precursors such as acetyl-CoA, hexanoyl-CoA, or olivetolic acid [28; 30], or involves supplementing the  
Figure 1: The proposed cannabinoid biosynthetic pathway, leading to the two major phytocannabinoids, THC and CBD. Enzymes are shown in blue. Bolded arrows represent an enzymatic reaction, whereas the dotted arrows represent non-enzymatic (heat or light) decarboxylation. media with sugars [31], such as galactose, to provide a more affordable alternative with a long series of gene stacking. For instance, the production of OA in _Dyctiostelium discoideum_ on a 300-L scale led to 4.8 ug/L [32]. OA production in _E. coli_ reached 80 mg/L after introducing a series of modifications to increase the CoAs pool [33]. However, the bacterial and yeast translation and post-translational mechanisms, as well as their metabolic chassis, are distinct from the mechanisms found in higher plants. This resulted in low yields, such as picomoles per unit of optical density, from these microorganisms when compared to the bioengineering and supplementation costs [31; 34]. By contrast, diatoms are photosynthetic organisms that share the main metabolic paths with plants, making them promising candidates for the production of heterologous compounds [9]. All the precursor pathways, such as acetyl-CoA, hexanoyl-CoA, GPP, etc., are present in diatom cells at different times of the cell cycle, which reduces the need to supplement the culture media with costly precursors or sugars [35]. Also, the extrachromosomal transformation of diatoms via bacterial conjugation allows the fairly stable expression of heterologous genes [36; 37]. Similar advances in the bioengineering of diatoms have been shown recently, allowing the production of monoterpenoids such as geraniol, an insect repellant, in the model diatom _Phaedactylum tricornutum_[36]. Thus, the aim of this study is to produce the CB precursor OA in the marine microalgae, _P. tricornutum_. Here, we inserted cannabis _Cs_TKS and _Cs_OAC genes into a stable optimized episome that was assembled in _S. cerevisiae_[38; 39]. The recombinant episome was then transformed via a trans-kingdom conjugation between _E. coli_ and _P. tricornutum_[38].