Abstract
The legalization of recreational and medicinal Cannabis sativa L. has been gaining global momentum, therefore increasing interest in Cannabis micropropagation as growers look for large-scale solutions to germplasm storage and clean plant propagation. Mother plants used in commercial propagation are susceptible to pests and disease and require considerable space. While micropropagation can produce disease-free starting material in less space, current published in vitro methods are not robust and few report high multiplication rates. Furthermore, these micropropagation methods rely on photoperiod-sensitive plants maintained in a perpetual vegetative state. Current methods are not adaptable to long-term tissue culture of day-neutral cultivars, which cannot be maintained in perpetual vegetative growth. In this study, we developed a micropropagation system which uses Cannabis inflorescences as starting materials. This study used two Cannabis cultivars, two plant growth regulators (PGR; 6-benzylaminopurine and meta-topolin) at different concentrations, and two different numbers of florets. Here, we show that floral reversion occurs from meristematic tissue in C. sativa florets and that it can be used to enhance multiplication rates compared to existing in vitro methods. Floret number was shown to have a significant impact on percent reversion, with pairs of florets reverting more frequently and producing healthier explants than single florets, while cultivar and PGR had no significant effect on percent reversion. Compared with our previously published nodal methods, the current floral reversion method produced up to eight times more explants per subculture. Floral reversion provides a foundation for effective inflorescence-based micropropagation systems in Cannabis.
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Acknowledgements
The authors gratefully acknowledge our industry partner, Hexo Corp. for the use of their plant material and Scott Golem for his help with greenhouse acclimatization of in vitro cultures. We would also like to thank Susan Lapos and the team at the Ontario Veterinary College’s Animal Health Laboratory for their histological expertise and support. The financial support of the Natural Sciences and Engineering Research Council of Canada is also gratefully acknowledged. Hexo Corp. and the Natural Sciences and Engineering Research Council of Canada were not involved in study design, data collection and analysis, the decision to publish or the preparation of the manuscript.
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This manuscript has been released as a pre-print at bioRxiv (Monthony et al. 2020b).
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AS Monthony has nothing to disclose; S Bagheri has nothing to disclose; Dr. Zheng has nothing to disclose; Dr. Jones reports grant funding though NSERC CRD grant (532081-18) and non-financial support from Hexo Inc. during the conduct of this study.
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Fig. S1
A sample spectrum of the LED lighting used to maintain reverting Cannabis sativa L. explants, obtained using a LI-COR LI-180 Spectrometer. PPFD: Photosynthetic Photon Flux Density. (PNG 6596 kb)
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Monthony, A.S., Bagheri, S., Zheng, Y. et al. Flower power: floral reversion as a viable alternative to nodal micropropagation in Cannabis sativa.. In Vitro Cell.Dev.Biol.-Plant 57, 1018–1030 (2021). https://doi.org/10.1007/s11627-021-10181-5
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DOI: https://doi.org/10.1007/s11627-021-10181-5