Medicinal Ganja

Conservation and Cultivation of Elite Varieties for Medicinal and Research

Applications

Ryan Francis (Presenter) Jodi Spence Gillian Rowe

3rd Biennial Conference on Science and Technology November 12, 2014

Outline

• Introduction

• In vitro Propagation of Cannabis sativa

• Hydroponic Propagation

• Greenhouse Propagation

• Conclusion

• References

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Introduction

• Cannabis sativa L. (Cannabaceace) originated in Asia, spread through Europe and later introduced into the America.

• Cannabis (Marijuana) is a dioecious annual herb.

• Produces equal number of male and female flower.

• This plant contains a unique class of terpenophenolic compounds called Cannabinoids.

Introduction

• More than 70 cannabinoids have been isolated from this herb (Lata et al 2009; Russo 2013).

• Main biologically active compounds are: Tetrahydro-cannabinol (THC) – Psychoactive.

• Cannabinol (CBD) – Nonpsycotophic (Mehmedic et al 2010).

Introduction

THC: Analgesic, anti-inflammatory, appetite stimulant, depression, rheumatism, fatigue, migrane and anti-emetic properties (Lata et al 2012).

CBD: Antimicrobial action, relieve anxiety, epilepsy, multiple sclerosis, dystonia and other neurological disorders (Clarke et al 1994).

• Pharmacological and therapeutic potential

Sources: http://jis.gov.jm/law-amended-facilitate-medical-ganja-industry/ http://moj.gov.jm/sites/default/files/laws/The%20Dangerous%20Drugs%20Act.pdf

Introduction

• Ganja (all parts of the Cannabis sativa plant) is one of the substances named in the Dangerous Drugs Act of 1948.

• It is an offense to cultivate, sell, deal in, transport, possess, import or export ganja.

• September 30, 2014 GOJ approved proposals for amendments to the Act to facilitate medicinal ganja and industrial hemp.

Introduction

• It is difficult to maintain pure strains of high THC or CBD yielding elite plants under field or greenhouse conditions.

• Plants are allogamous (cross-fertilization).

• Difficult to maintain elite cultivars by seed.

Breeding and Conservation challenges

In Vitro Propagation of Cannabis sativa

Advantages of Tissue Culture Propagation

• Better storage method than seeds, which can be highly variable and produce less efficacious genotypes (Meijer et al 1992).

• Facilitate the conservation of elite cultivars of Cannabis sativa.

Tissue Culture of Cannabis sp.

Sterilization

• Surface sterilize with 0.5% NaOCl (15% v/v bleach) and 0.1 % Tween 20 for 20 mins.

• 3 five mins rinses in sterile distilled water.

Materials and Methods

Plant material/explant

• Nodal segments with axillary buds (approx. 1cm)

• Obtained from healthy branches of high yielding Cannabis sp.

Lata et al 2009

Tissue Culture

Supplemented with: • 3% (w/v) sucrose • 0.8% (w/v) agar • 0.5 μM thidiazuron (TDZ) • pH 5.7

Lata et al 2009b

Initiation

Disinfected explants transferred to baby food jars containing 25 ml of sterile Murashige and Skoog’s (MS) medium (Murashige and skoog 1992).

Tissue Culture

Multiplication

4-6 weeks after initiation emerging shoots should be transferred to MS multiplication medium supplemented with:

Lata et al 2009b

• 0.5 μM TDZ

• 7.0 μM Gibberellic acid (GA3)

Tissue Culture Rooting

This should be done on medium containing

Lata et al 2009b

• ½ strength MS medium, supplemented with

• 500 mg/L activated charcoal

• 5.0 μM Indole-3-butyric acid (IBA)

Tissue Culture

Acclimatization

• Rooted plants should be transferred to Jiffy pots containing potting mix.

Lata et al 2009b

• New growth after 2-3 weeks.

Summary

Adopted from Lata et al 2009b

Hydroponic Cultivation

What is hydroponic cultivation?

•Hydroponics is a method of growing

plants using mineral nutrient solutions

in water, without soil (Douglas 1985).

•Terrestrial plants may be grown with

their roots in the mineral nutrient

solution only or in an inert medium,

such as perlite, gravel, coco peat and

others.

Benefits of Hydroponics Over Soil

• No soil required.

• Lower water requirement as the water can be reused.

• Allows full control over the plant nutrition levels.

• No fertilizer run off is released into the environment.

• Stable and high yields.

• Reduces soil-borne pests and diseases.

• Ease of harvesting.

• The pH of nutrient solution can be controlled.

Disadvantages of Hydroponics

• Requires more knowledge and expertise.

• System problems lead to rapid crop death.

• Requires constant monitoring.

• High humidity favours some pathogens.

• High maintenance costs for indoor systems.

Types of Hydroponic Systems

• Static solution

• Continuous flow

• Ebb and flow

• Run to waste

• Aeroponics and fogponics

• Passive sub-irrigation

• Deep water culture

Types of Hydroponic Systems

Source: http://www.nosoilsolutions.com/6-different-types-hydroponic-systems/

Types of Substrates

• Sand

• Expanded clay aggregate

• Coco peat

• Gravel

• Brick shards

• Perlite

• Rockwool

• Growstones

• Rice hulls

• Vermiculite

• Pumice

Perlite Growstones Rockwool

Expanded clay Coco coir

Pumice

Source: http://ezgrogarden.com/growing-organic/grorock-hydroponic-substrate/

Growing in a Greenhouse

Greenhouse

Purpose: It heats up because incoming visible sunshine is absorbed inside the structure. Air warmed by the heat from warmed interior surfaces is retained in the building by the roof and wall; the air that is warmed near the ground is prevented from rising indefinitely and flowing away.

A greenhouse is a structural building with different types of covering materials, such as a glass or plastic roof and frequently glass or plastic walls.

Growing Cannabis sp. in a Greenhouse

A Cannabis sp. specific greenhouse typically has a blackout system to adjust the photoperiod. There are two phases of the greenhouse:

Vegetative Phase - The vegetative room offers plants 18 hours of light and 6 hours of dark. The plants will grow forever. They grow like trees. You can keep strains alive for decades. Once the plants reach a desirable size, they are moved to the flowering phase of the greenhouse.

Flowering Phase – To induce flowering blackout curtains are used to maintain 12 hours light and 12 hours dark. In 4-8 weeks flowers ripen. (Grotenhermen and Russo,2002).

Advantages of Greenhouse

• Ability to manipulate environment

• Protection

a. Wind and pests

b. Extremely hot weather

c. Cross-pollination

d. Excess water/moisture

From:

Source: http://www.maryjanesgarden.com/growing/greenhouse-growing

Advantages of Greenhouse

• Plant disease suppression.

• Avoid workers exposing to severe weather conditions.

• Structure with controlled access and security features can be installed.

Safety

Advantages of Greenhouse

Improve Growth

• Propagate new plants.

• Increase yields and quality.

Source: http://www.usatoday.com/story/news/nation/2014/08/12/colorado-marijuana-sales/13915859/

Disadvantages of Greenhouse

• Construction Costs

• High Energy/ Operation Costs

• Maintenance

• Tools and supplies – soil, pots etc.

• Human Resources

Conclusion

• Tissue culture can be used regenerate a large number of Cannabis sativa plants in a short period of time (Slusarkiewicz-Jarzina et al 2005, Lata et al 2009a).

• Hydroponic cultivation of marijuana offers an alternative to soil and has many benefits over traditional growing methods (Chandra et al 2010).

• Tissue culture can be used to conserve elite cultivars of Cannabis sativa in vitro (Lata et al 2009b;2012).

• Growing marijuana in a greenhouse allows for control over growing conditions in a protected environment (Chandra et al 2011).

• These techniques can be used for germplasm exchange between laboratories and in breeding to produce high yielding varieties for the pharmaceutical industry.

References Angus, G. (2014, September 30). Law to be Amended to Facilitate Medical Ganja Industry. Jamaica Information Service. Retrieved from http://jis.gov.jm/law-amended-facilitate-medical-ganja-industry/. Chandra, S., Lata, H., Khan, I. A., & ElSohly, M. A. (2010). Propagation of elite Cannabis sativa for the production of D9-Tetrahydrocannabinol (THC) using biotechnological tools. Medicinal plant biotechnology, 98-114.

Chandra, S., Lata, H., Khan, I. A., & ElSohly, M. A. (2011). Temperature response of photosynthesis in different drug and fiber varieties of Cannabis sativa L. Physiology and Molecular Biology of Plants, 17(3), 297-303. Clarke, R. C., & Pate, D. W. (1994). Medical marijuana. J. International Hemp Asso, 9-12. Colorado firm partners with Jamaicans to set up medical ganja lab on island. (2014, October 5). The Jamaica Observer. Retrieved from http://www.jamaicaobserver.com/business/Colorado-firm-partners-with-Jamaicans--to-set-up-medical-ganja-lab-on-island_17680298. Douglas, J. S. (1985). Advanced guide to hydroponics (No. new edition). Pelham Books. Dorm, D. (September 26, 2013). Greenhouse Growing For Efficient Medical Marijuana Production. Retrieved from http://www.medicaljane.com/2013/09/26/greenhouse-growing-efficient-medical-marijuana-production/. Dreier, F. (March 31, 2014). Greenhouse Technology Shakes Up Economics of Cannabis Cultivation. In Marijuana Business Daily. Retrieved from http://mmjbusinessdaily.com/the-basics-of-marijuana-greenhouse-technology/. Espiritu, K. (2011). Hydroponic Media. Retrieved October 13, 2014, from http://www.epicgardening.com/hydroponic-growing-media/.

Grotenhermen, F. and Russo, E. (2002). Cannabis and Cannabinoids: Pharmacology, Toxicology, and Therapeutic Potential.

Binghampton , N.Y. Haworth Integrative Healing Press.

References

Growing Cannabis in a Greenhouse. (n.d.). Retrieved from http://www.dutch-passion.nl/en/grow-info/growing-cannabis-in-a-greenhouse/. Haze, N. (n.d.). Soil vs. Hydroponics. Retrieved October 2, 2014, from http://www.growweedeasy.com/marijuana-soil-vs-hydroponics. How to Grow Marijuana Hydroponically. (n.d.). Retrieved October 3, 2014, from http://www.wikihow.com/Grow-Marijuana-Hydroponically How to Grow Top Shelf Cannabis Using Greenhouses. (n.d.). Retrieved from http://www.triqsystems.com/resources/grow-top-shelf-cannabis.html. Hydroponics. (n.d.) Retrieved October 3, 2014, from http://en.wikipedia.org/wiki/Hydroponics. Lata, H., Chandra, S., Khan, I. A., & ElSohly, M. A. (2009a). Propagation through alginate encapsulation of axillary buds of Cannabis sativa L.—an important medicinal plant. Physiology and Molecular Biology of Plants, 15(1), 79-86. Lata, H., Chandra, S., Khan, I., & ElSohly, M. A. (2009b). Thidiazuron-induced high-frequency direct shoot organogenesis of Cannabis sativa L. In Vitro Cellular & Developmental Biology-Plant, 45(1), 12-19. Lata, H., Chandra, S., Mehmedic, Z., Khan, I. A., & ElSohly, M. A. (2012). In vitro germplasm conservation of high Δ9-tetrahydrocannabinol yielding elite clones of Cannabis sativa L. under slow growth conditions. Acta Physiologiae Plantarum, 34(2), 743-750.

References Mehmedic, Z., Chandra, S., Slade, D., Denham, H., Foster, S., Patel, A. S., & ElSohly, M. A. (2010). Potency Trends of Δ9‐THC and Other Cannabinoids in Confiscated Cannabis Preparations from 1993 to 2008*. Journal of Forensic Sciences, 55(5), 1209-1217.

Murashige, T and Skoog, F. (192). A revised medium for rapid growth and bioassays with tobacco tissue culture. Physiologia Plantarum 15: 473-497.

Neita, L. (2014, October 5). The Greening of Jamaica. The Jamaica Observer. Retrieved from http://www.jamaicaobserver.com/columns/The-greening-of-Jamaica_17676447.

Quimby M. 1974. Botany of Cannabis sativa. In: Mateos-Gomez JL (ed) Archivos be investigacion medica/ El Instituto Mexicano del Seruto Social.

Russo, E. B. (2013). Cannabis and cannabinoids: pharmacology, toxicology, and therapeutic potential. Routledge.

Slusarkiewicz-Jarzina, A. U. R. E. L. I. A., Ponitka, A., & 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.

THANK YOU!

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