Improvement of mineral nutrition and rooting efficiency of Cannabis sativa L. for in vitro large-scale propagation

Cannabis micropropagation is an effective method for producing clean plants in the new and expanding cannabis industry. However, existing protocols are not efficient enough to mass propagate for industry demands and to preserve large numbers of elite cultivars. This research was designed to validate optimal mineral nutrition for cannabis micropropagation. Currently, Murashige and Skoog (MS) basal salts are used in commercially available cannabis medium. Three mineral nutrient factors were defined from MS salts: ammonium nitrate, potassium nitrate, and mesonutrients (CaCl₂·2H₂O, KH₂PO₄, MgSO₄·7H₂O). A total of 20 treatments (MS1-20) were designed, and each factor was varied over a range of concentrations. Leaf canopy area, shoot fresh weight, and multiplication number were measured to quantify the effects of mineral nutrients on the in vitro growth of three cannabis cultivars. All cultivars responded positively to increased nitrogen and mesonutrients (MS6) relative to MS medium. Plantlets grown on MS6 exhibited a significant increase in canopy area and fresh weight in two out of three studied cultivars. Additionally, an increase in multiplication number was noted on MS6 with an average increase in shoots per explant of 52% across three cultivars. The addition of potassium silicate (4.86 mM) to MS6 media reduced hyperhydricity symptoms in three hyperhydric cultivars. An improved root induction was observed on inert media, rockwool, and Oasis foam plugs when compared to standard culture media (½ MS containing 2.4 µM IBA and MS containing 2 µM m-topolin) that are routinely used as cannabis rooting media. Plantlets grown in rockwool resulted in healthy shoots and a high root fresh weight after 4 to 6 wk with mineral nutrient solution while plantlets grown on standard culture media did not produce roots during this period. Together, this study presents an end-to-end efficient procedure for cannabis micropropagation and preservation to be used in industrial large-scale production.