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Propagation through alginate encapsulation of axillary buds of Cannabis sativa L. — an important medicinal plant

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Abstract

Cannabis sativa L. (Cannabaceae) is an important medicinal plant well known for its pharmacologic and therapeutic potency. Because of allogamous nature of this species, it is difficult to maintain its potency and efficacy if grown from the seeds. Therefore, chemical profile-based screening, selection of high yielding elite clones and their propagation using biotechnological tools is the most suitable way to maintain their genetic lines. In this regard, we report a simple and efficient method for the in vitro propagation of a screened and selected high yielding drug type variety of Cannabis sativa, MX-1 using synthetic seed technology. Axillary buds of Cannabis sativa isolated from aseptic multiple shoot cultures were successfully encapsulated in calcium alginate beads. The best gel complexation was achieved using 5 % sodium alginate with 50 mM CaCl2.2H2O. Regrowth and conversion after encapsulation was evaluated both under in vitro and in vivo conditions on different planting substrates. The addition of antimicrobial substance — Plant Preservative Mixture (PPM) had a positive effect on overall plantlet development. Encapsulated explants exhibited the best regrowth and conversion frequency on Murashige and Skoog medium supplemented with thidiazuron (TDZ 0.5 μM) and PPM (0.075 %) under in vitro conditions. Under in vivo conditions, 100 % conversion of encapsulated explants was obtained on 1:1 potting mix- fertilome with coco natural growth medium, moistened with full strength MS medium without TDZ, supplemented with 3 % sucrose and 0.5 % PPM. Plantlets regenerated from the encapsulated explants were hardened off and successfully transferred to the soil. These plants are selected to be used in mass cultivation for the production of biomass as a starting material for the isolation of THC as a bulk active pharmaceutical.

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Abbreviations

CaCl2.2H2O:

Calcium chloride

MS:

Murashige and Skoog

PGR:

Plant Growth Regulator

PPM:

Plant Preservative Mixture

TDZ:

Thidiazuron

References

  • Abrams DI, Jay CA, Shade SB, Vizoso H, Reda H, Press S, Kelly ME, Rowbotham MC and Petersen KL (2007). Cannabis in painful HIV-associated sensory neuropathy: A randomized placebo-controlled trial. Neurol 68: 515–521.

    Article  CAS  Google Scholar 

  • Adriani M, Piccioni E and Standardi A (2000). Effect of different treatments on the conversion of ‘Hayward’ kiwifruit synthetic seeds to whole plants following encapsulation of in vitro-derived buds. N Z J Crop Horti Sci 28: 59–67.

    Google Scholar 

  • Bapat VA, Mhatre M and Rao PS (1987). Propagation of Morus indica L. (mulberry) by encapsulated shoot buds. Plant Cell Rep 6: 393–395.

    Article  Google Scholar 

  • Bapat VA and Rao PS (1990). In vitro growth of encapsulated axillary buds of mulberry (Morus indica L.). Plant Cell Tiss Org Cult 20: 69–70.

    Article  Google Scholar 

  • Bing X, Ning L, Jinfeng T and Nan G (2007). Rapid tissue culture method of Cannabis sativa for industrial uses. CN 1887043 A 20070103 Patent, p 9.

  • Bornman CH (1993). Maturation of Somatic Embryos. In: Synseeds: Application of Synthetic Seeds to Crop Improvement (Ed. Redenbaugh K.), CRC Press, Boca Raton, Florida, pp. 105–114.

    Google Scholar 

  • Brenneisen R, Egli A, Elsohly MA, Henn V and Spiess Y (1996). The effect of orally and rectally administered -tetrahydrocannabinol on spasticity. A pilot study with two patients. Int J Clin Pharm Ther 34(1): 446.

    CAS  Google Scholar 

  • Brischia R, Piccioni E and Standardi A (2002). Micropropagation and synthetic seed in M.26 apple rootstock (II): A new protocol for production of encapsulated differentiating propagules. Plant Cell Tiss Org Cult 68: 137–141.

    Article  Google Scholar 

  • Castillo B, Smith MAL and Yadava UL (1998). Plant regeneration from encapsulated somatic embryos of Carica papaya L. Plant Cell Rep 17: 172–176.

    Article  CAS  Google Scholar 

  • Duke JA and Wain KK (1981). Medicinal Plants of the world, Computer index with more than 85.000 entries In: Handbook of Medicinal Herbs (Ed. Duke J.A.), CRC press, Boca Raton, Florida, pp. 96.

    Google Scholar 

  • Fowke LC, Attree SM and Pometry MK (1994). Production of vigorous desiccation-tolerant white spruce (Picea glauca {Moench} Voss.) synthetic seeds in a bioreactor. Plant Cell Rep 13: 601–606.

    Article  Google Scholar 

  • Fujii JA, Slade D and Redenbaugh K (1993). Planting of artificial seeds and somatic embryos. In: Synseeds: Application of Synthetic Seeds to Crop Improvement (Ed. Redenbaugh K.), CRC Press, Boca Raton, Florida, pp. 183–202.

    Google Scholar 

  • Ganapathi TR, Suprasanna P, Bapat VA and Rao PS (1992). Propagation of banana through encapsulated shoot tips. Plant Cell Rep 11: 571–575.

    Article  Google Scholar 

  • Ganapathi TR, Srinivas I, Suprasanna P and Bapat VA (2001). Regeneration of plants from alginated-encapsulated somatic embryos of banana cv. Rasthali (Musa spp. AAB group). Biol Plant 37: 178–181.

    CAS  Google Scholar 

  • Grinspoon L and Bakalar JB (1995). Marihuana as medicine. JAMA 273: 1875–1876.

    Article  PubMed  CAS  Google Scholar 

  • Guri AZ and Patel KN (1998). Compositions and methods to prevent microbial contamination of plant tissue culture media. United States Patent 5: 750, 402.

    Google Scholar 

  • Hao YJ and Deng XX (2003). Genetically stable regeneration of apple plants from slow growth. Plant Cell Tiss Org Cult 72: 253–260.

    Article  CAS  Google Scholar 

  • Larkin PJ, Davies PA and Tanner GJ (1988). Nurse culture of low number of Medicago and Nicotiana protoplasts using calcium alginate beads. Plant. Sci. 58: 203–210.

    Article  CAS  Google Scholar 

  • Lata H, Chandra S, Khan I and ElSohly MA (2008). Thidiazuron induced high frequency direct shoot organogenesis of Cannabis sativa L. In vitro Cell Dev Biol Plant (accepted).

  • Long LE, Malone DT and Taylor DA (2005). The pharmacological actions of cannabidiol. Drugs of the Future 30(7): 747.

    Article  CAS  Google Scholar 

  • Mandolino G and Ranalli P (1999). Advances in biotechnological approaches for hemp breeding and industry. In: Advances in hemp research (Ed. Ranalli P.), Haworth Press, New York, pp. 185–208.

    Google Scholar 

  • Mathur J, Ahuja PS, Lal N and Mathur AK (1989). Propagation of Valeriana wallichii DC using encapsulated apical and axial shoot buds. Plant Sci 60: 111–6.

    Article  Google Scholar 

  • Mattes RD, Egelman K, Shaw LM and Elsohly MA (1994). Cannabinoids appetite stimulation. Pharmacol Biochem Behav 44(3): 745–747.

    Article  Google Scholar 

  • Mechoulam S, Lander N, Dikstein S, Carlini EA and Blumenthal M (1976). On the Therapeutic Possibilities of Some Cannabinoids. In: The Therapeutic Potential of Marihuana (Eds. Cohen, S. and Stillman, R) Plenum Press, New York pp. 36.

    Google Scholar 

  • Micheli M, Pellegrino S, Piccioni E and Standardi A (2002). Effects of double encapsulation and coating on synthetic seed conversion in M.26 apple rootstock. J. Microencap 19(3): 347–356.

    Article  CAS  Google Scholar 

  • Micheli M, Hafiz IA and Standardi A (2007). Encapsulation of in vitro-derived explants of olive (Olea europaea L. cv. Moraiolo) II Effects of storage on capsule and derived shoots performance. Sci Horti 113: 286–292.

    Article  Google Scholar 

  • Murashige T and Skoog F (1962). A revised medium for rapid growth and bioassays with tobacco tissue cultures. Physiol Plant 15: 473–497.

    Article  CAS  Google Scholar 

  • Onishi N, Sakamoto Y and Hirosawa T (1994). Synthetic seeds as an application of mass production of somatic embryos. Plant Cell Tiss Org Cult 39: 137–145.

    Article  Google Scholar 

  • Pattnaik S and Chand PK (2000). Morphogenic response of the alginate encapsulated axillary buds from in vitro shoot cultures of six mulberries. Plant Cell Tiss Org Cult 64: 177–185.

    Article  Google Scholar 

  • Piccioni E and Standardi A (1995). Encapsulation of micropropagated buds of six woody species. Plant Cell Tiss Org Cult 42: 221–226.

    Article  Google Scholar 

  • Pryce G and Baker D (2005). Emerging properties of cannabinoid medicines in management of multiple sclerosis. Trends in Neurosci 28(5): 272–276.

    Article  CAS  Google Scholar 

  • Richez-Dumanois C, Braut-Boucher F, Cosson L and Paris M (1986). Multiplication vegetative in vitro du chanvre (Cannabis sativa L.) Application a la conservation des clones selectiones. Agronomie 6: 487–495.

    Article  Google Scholar 

  • SAS Institute, Inc. (2003). Statistical Analysis Systems User’s guide: Statistics version 9.1. SAS Institute, Cary, NC.

    Google Scholar 

  • Sharma TR, Singh BM and Chauhan RS (1994). Production of disease free encapsulated buds of Zingiber officinale Rose. Plant Cell Rep 13: 300–302.

    Article  CAS  Google Scholar 

  • Sirikantaramas S, Taura F, Morimoto S and Shoyama Y (2007). Recent Advances in Cannabis sativa Research: Biosynthetic Studies and Its Potential in Biotechnology. Curr Pharma Biotechnol 8(4): 237–243.

    Article  CAS  Google Scholar 

  • Slusarkiewicz-Jarzina A, Ponitka A and Kaczmarek Z (2005). Influence of cultivar, explant source and plant growth regulator on callus induction and plant regeneration of Cannabis sativa L. Acta Biol Craco Series Bot 47(2): 145–151.

    Google Scholar 

  • Small E and Marcus D (2002). Hemp: A new crop with new uses for North America. In: Trends in new crops and new uses (Eds. Janick, J. and Whipkey, A.), ASHS Press, Alexandria, VA, pp. 284–326.

    Google Scholar 

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Authors and Affiliations

  1. National Center for Natural Products Research, School of Pharmacy, The University of Mississippi, University, MS, 38677, USA

    Hemant Lata, Suman Chandra, Ikhlas A. Khan & Mahmoud A. ElSohly

  2. Department of Pharmacognosy, School of Pharmacy, The University of Mississippi, University, MS, 38677, USA

    Ikhlas A. Khan

  3. Department of Pharmaceutics, School of Pharmacy, The University of Mississippi, University, MS, 38677, USA

    Mahmoud A. ElSohly

Authors
  1. Hemant Lata
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  2. Suman Chandra
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  3. Ikhlas A. Khan
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  4. Mahmoud A. ElSohly
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Correspondence to Hemant Lata.

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Lata, H., Chandra, S., Khan, I.A. et al. Propagation through alginate encapsulation of axillary buds of Cannabis sativa L. — an important medicinal plant. Physiol Mol Biol Plants 15, 79–86 (2009). https://doi.org/10.1007/s12298-009-0008-8

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  • DOI: https://doi.org/10.1007/s12298-009-0008-8

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