# Phytocannabinoids Biosynthesis in Angiosperms, Fungi, and Liverworts and Their Versatile Role
## 5 Biotechnology and In Vitro Propagation of Cannabis  
Cannabis is an essential plant as it yields phytocannabinoid, which has multiple applications. Cannabis cultivation is regulated in several countries, so alternative biotechnological and in vitro tissue culture approaches require attention. Meanwhile, these approaches are beneficial to preserve cultivars or clones of plants with specific metabolites such as phytocannabinoid. Micropropagation generates genetically stable plants; thus, this method is beneficial for the clonal multiplication of cannabis. Nevertheless, micropropagation of the cannabis plant is performed via adventitious buds and axillary buds located on the nodal segment. Another method that is synthetic seed technology (encapsulation of axillary bud or nodal segment in calcium alginate seeds) has also gained importance in plant clonal propagation. It displays homogenous growth and development of plants and genetic stability even after long storage [65; 66]. Cannabis is not a well-known recalcitrant plant for transformation. The plant's regeneration capacity is low and is completely dependent on plant tissue, age, explants, and a combination of plant growth regulators (PGRs), i.e., indole-3-acetic acid (IAA), indole-3-butyric acids, 1-naphthaleneacetic acid (NAA), 2,4-dichlorophenoxyoxyacetic acid, and kinetin. Moreover, successful transformation is performed via _Agrobacteriumium tumefaciens_; cells that do not regenerate the shoots are undifferentiated and can be transformed via phosphomannose isomerize and calorimetric assays, which showed efficacious expression of the transgene [67]. Cannabis transformation using shoot tip explants regenerates a high amount of cannabinoid after infection with _Agrobacteriumium tumefaciens_[68; 69]. For the successful regeneration of cannabis, thidiazuron (behavior as cytokinin-like compound) with kinetin (represent cytokinns) has not only the effect on the shoots but also shows a high yielding capacity of phytocannabinoids. Dicamba (3,6-dichloro-2-methoxybenzoic acid) herbicide is also known for inducing the regeneration of shoots from calli. Moreover, a most potent and efficient transformation of cannabis was carried out using 1-2 cm of hypocotyl explants and by supplementation of zeatin and 6-benzylaminopurine [65]. Another important tissue culture technique used for the production of bioactive cannabinoids showing pharmaceutical effects is hairy root culture. In this system, both _Agrobacteriumium tumefaciens_ and _Agrobacteriumium rhizgenes_ are used for transformation to yield cannabinoids by establishing a hairy root system. Meanwhile, the most responsive tissue for infection in this system is hypocotyl, which produces large quantities of hypocotyls or engineered plants for the production of bioactive compounds that are industrially valuable; for example, \(\Delta^{9}\)-THCA is produced by expressing \(\Delta^{9}\)-THCAS. The hairy root technique is obtained by the increased growth rate, independent of a hormone, and has the same metabolic potential as the original plant organ. In the hairy root system, cannabis callus was cultured on a full-strength B5 medium feed with an NAA or IAA displayed increase in the accumulation of phytocannabinoids. Rhizenesis induction in cannabis undifferentiated cells is crucial, as it can be performed on calli overexpressing some important transcription factors (TFs) and genes which are involved in cannabinoid synthesis. Additionally, cannabis hairy root culture is an efficient method implemented with adsorbents to reduce toxicity problems [65; 67]. Cannabis cell suspension culture transformation with the genes involved in the cannabinoid synthesis pathway offers a great opportunity to enhance bioactive phyto-cannabinoid production possessing pharmacological potential. Additionally, increased production via cannabis cell suspension culture can be obtained through TF.