Ecophysiological Aspects of in vitro Propagated Cannabis sativa L. Plants During Acclimatization

Acclimatization is an important final step in the successful completion of a micropropagation protocol. Since plants produced in nutrient rich substrate under sterile conditions are fragile, the transition during hardening from in vitro to an ex vitro environment can be very critical, and subsequently affect the field performance of the plants. Poor survival rates of in vitro plantlets after ex vitro transfer can limit the application of a micropropagation technique. Plant development under in vitro conditions can result in reduced leaf epicuticular wax, low photosynthetic capacity, increased stomatal density and deficient vascular connection between the roots and shoots. However, during this acclimatization or hardening, plants undergo physiological and morphological changes that improve their ability to grow in nonsterile environment.

This work describes the photosynthetic and water vapor exchange performance of micropropagated plants of Cannabis sativa and their comparison with vegetatively grown plants from the same mother plant during acclimatization under climatic controlled grow room conditions. Well rooted cultures were carefully taken out of the culture medium, washed thoroughly and pre-incubated in coco natural growth medium in small jiffy pots for 10 days at 25 ± 3 ^oC and 18 hours under cool florescent lights. The pots were covered with polyethylene bags to maintain humidity. These plants were then transferred in sterile potting mix in larger pots. All these plantlets were kept under controlled environmental conditions with vegetatively grown plants from the same mother plant in the indoor cultivation facility at the Coy-Waller laboratory, The University of Mississippi. Tissue culture raised plants were then evaluated for their ecophysiological characteristics {net photosynthesis (P _N), transpiration (T _R), stomatal conductance (gCO)₂, intercellular CO₂ concentration (C _i), ratio of intercellular CO₂ concentration to ambient CO₂ concentration (C _i/C _a) and water use efficiency (WUE)} and compared with the vegetatively grown plants from the same mother plant. Our results reveal no statistically significant differences in groß morphology and ecophysiological characteristics of the two types of plants and thus, confirm the clonal fidelity of micropropagated plants.

Acknowledgements: This work was supported in part by the National Institute on Drug Abuse (NIDA), National Institutes of Health (NIH), Department of Health and Human Services, USA, Contract No. N01DA-10 – 7773.