Non-thermal plasma improved callogenesis performance and elicited the production of cannabinoids by modifying DNA methylome, expression of WRKY1 and ERF1B transcription factors, and expression of genes that contributed to the biosynthesis of cannabinoids.

The current decade has witnessed notable advancement towards the utilization of non-thermal (cold) plasma in multidisciplinary fields such as plant sciences. This study intends to validate whether cold plasma contributes to improving callogenesis performance and eliciting the production of cannabinoids in cannabis. The cannabis-derived calli were treated with plasma at different exposure times, including 0, 60, 120, and 180 s. The plasma priming improved the callogenesis performance and callus biomass by an average of 46.6%. The molecular assessment (MSAP method) validated how the plasma priming is epigenetically associated with variation in DNA methylome in the cannabis calli. The cold plasma treatments transcriptionally upregulated the expression of WRKY1 and ERF1B transcription factors by averages of 3.5- and 3.8-fold. The plasma treatment also stimulated the transcription of OLS, OAC, CBGAS, CBDAS, and THCAS genes involved in the biosynthesis of cannabinoids. The HPLC assessment proved the high potency of cold plasma to enhance the synthesis of cannabinoids, including Cannabigerol (CBG), Cannabidiol (CBD), and cannabinol (CBN). The plasma-primed calli contained higher concentrations of proteins (56%), proline (38%), and soluble phenols (40%). The activities of peroxidase and catalase enzymes showed a similar upward trend in response to the plasma. The profound increase in the concentrations of soluble sugars resulted from the plasma treatments. The plasma priming of calli contributed to the significant upregulation in the activity of the phenylalanine ammonia-lyase enzyme. This biological assessment study validates the high potency of plasma priming to elicit the biosynthesis of cannabinoids in cannabis calli.