Abstract
Cannabis sativa L. (Marijuana; Cannabaceae), one of the oldest medicinal plants in the world, has been used throughout history for fiber, food, as well as for its psychoactive properties. The dioecious and allogamous nature of C. sativa is the major constraint to maintain the consistency in chemical profile and overall efficacy if grown from seed. Therefore, the present optimized in vitro propagation protocol of the selected elite germplasm via direct organogenesis and quality assurance protocols using genetic and chemical profiling provide an ideal pathway for ensuring the efficacy of micropropagated Cannabis sativa germplasm. A high frequency shoot organogenesis of C. sativa was obtained from nodal segments in 0.5 μM thidiazuron medium and 95 % in vitro rhizogenesis is obtained on half-strength MS medium supplemented with 500 mg/L activated charcoal and 2.5 μM indole-3-butyric acid. Inter Simple Sequence Repeats (ISSR) and Gas Chromatography-Flame Ionization Detection (GC-FID) are successfully used to monitor the genetic stability in micropropagated plants up to 30 passages in culture and hardened in soil for 8 months.
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References
Hammond CT, Mahlberg PG (1977) Morphogenesis of capitate glandular hairs of Cannabis sativa (Cannabaceae). Am J Bot 64:1023–1031
ElSohly MA, Gul W (2014) Constituents of Cannabis sativa. In: Pertwee RG (ed) Handbook of Cannabis. Oxford University Press, Oxford, pp 3–22
Sirikantaramas S, Taura F, Morimoto S, Shoyama Y (2007) Recent advances in Cannabis sativa research: biosynthetic studies and its potential in biotechnology. Curr Pharm Biotechnol 8:237–243
Faisal M, Anis M (2002) Rapid in vitro propagation of Rauvolfia tetraphylla L.—an endangered medicinal plant. Physiol Mol Biol Plants 8(2):295–299
Patial V, Devi K, Sharma M, Bhattacharya A, Ahuja PS (2012) Propagation of Picrorhiza kurroa Royle ex Benth: an important medicinal plant of Western Himalaya. J Med Plant Res 6(34):4848–4860
Sivanesan I (2007) Direct regeneration from apical bud explants of Withania somnifera Dunal. Indian J Biotechnol 6:125–127
Husain MK, Anis M, Shahzad A (2007) In vitro propagation of Indian kino (Pterocarpus marsupium Roxb.) using Thidiazuron. In Vitro Cell Dev Biol Plant 43:59–64
Shahzad A, Ahmad N, Anis M (2006) An improved method of organogenesis from cotyledonary callus of Acacia sinuate (Lour.) Merr. using Thidiazuron. J Plant Biotechnol 8:15–19
Lata H, Bedir E, Hosick A, Ganzera M, Khan IA, Moraes RM (2002) In vitro plant regeneration from leaf-derived callus in Black cohosh (Cimicifuga racemosa). Planta Med 68:912–915
Shahzad A, Parveen S, Fatema M (2010) Development of a regeneration system via nodal segment culture in Veronica anagallis-aquatica L.—an amphibious medicinal plant. J Plant Interact 5:115–121
Lata H, Chandra S, Khan IA, ElSohly MA (2009) Thidiazuron induced high frequency direct shoot organogenesis of Cannabis sativa L. In Vitro Cell Dev Biol Plant 45:12–19
Loh WHT, Hartsel SC, Robertson W (1983) Tissue culture of Cannabis sativa L. and in vitro biotransformation of phenolics. Z Pflanzenphsiol: 395–400
Richez-Dumanois C, Braut-Boucher F, Cosson L, Paris M (1986) Multiplication vegetative in vitro du chanvre (Cannabis sativa L.). Application a la conservation des clones selectionnes. Agronomie 6:487–495
Mandolino G, Ranalli P (1999) Advances in biotechnological approaches for hemp breeding and industry. In: Ranalli P (ed) Advances in hemp research. Haworth Press, New York, pp 185–208
Slusarkiewicz-Jarzina A, Ponitka A, Kaczmarek Z (2005) Influence of cultivar, explant source and plant growth regulator on callus induction and plant regeneration of C annabis sativa L. Acta Biol Cracov Bot 47(2):145–151
Bing X, Ning L, Jinfeng T, Nan G (2007) Rapid tissue culture method of Cannabis sativa for industrial uses. CN 1887043 A 20070103 Patent. pp 9
Murashige T, Skoog F (1962) A revised medium for rapid growth and bioassays with tobacco tissue cultures. Physiol Plant 15:473–497
Ross SA, Parker M, Arafat R, Lovett K, ElSohly MA (1996) The analysis of confiscated marijuana samples for different cannabinoids using GC/FID. Am Lab:16–17
Acknowledgements
This work was supported in part through the National Institute on Drug Abuse (NIDA), National Institute of Health (NIH), Department of Health and Human Services, USA, contract no. N01DA-10-7773.
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Lata, H., Chandra, S., Khan, I.A., ElSohly, M.A. (2016). In Vitro Propagation of Cannabis sativa L. and Evaluation of Regenerated Plants for Genetic Fidelity and Cannabinoids Content for Quality Assurance. In: Jain, S. (eds) Protocols for In Vitro Cultures and Secondary Metabolite Analysis of Aromatic and Medicinal Plants, Second Edition. Methods in Molecular Biology, vol 1391. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-3332-7_19
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DOI: https://doi.org/10.1007/978-1-4939-3332-7_19
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