The Society of In Vitro Biology (SIVB) has been interested in the tissue culture of important plants since its inception. Indeed, the society began as the Tissue Culture Association (TCA) in 1949. Cannabis research has been presented at the SIVB meetings in 2017 and 2018, while cannabis articles have been written in In Vitro Cellular & Developmental Biology – Plant, the publishing arm of the Society. At the June 2019 meeting of SIVB, there were 12 presentations, these are summarized here. More details can be obtained from the publication:
In Vitro Cellular & Developmental Biology – Animal, Volume 55, abstract issue and In Vitro Cellular & Developmental Biology – Plant, Volume 55, issue 4.
Cataloguing existing variation and rebuilding better Cannabis genomes for new markets. John McKay. (P-35, S23)
The cannabis (hemp) breeding program at New West Genetics was explained.
Polyploidization for the genetic improvement of Cannabis sativa. Jessica Parsons. (P-36, S24)
Polyploidization has been used to develop hemp-type strains. Now it is being explored for drug-type strains. Tetraploid drug-type lines developed using oryzalin showed significant differences in leaf and stomata size, glandular density and exhibited changed terpene profiles.
The role of genetics on large-scale hemp production. Jonathan Vaught. (P-37, S24)
The importance of genomics-driven breeding to produce hemp varieties that are reliable, consistent and profitable was explained.
Embryo excision of Cannabis sativa L. Kyle Chabrier, H Jones. (P-3025, pg 490)
Due to the past legal status of Cannabis, much of its growth and production has gone widely unstudied and unregulated thus research has to start at the beginning. Embryo excision is a technique useful for determining seed viability, overcoming seed dormancy, obtaining plants of varied ploidy, shortening the breeding cycle and allowing circumvention of incompatible, intergeneric hybridization. Embryos excised aseptically all germinated. These developed rapidly without the presence of any outside hormonal additives and could be subcultured.
Development of tissue culture methods for marijuana (Cannabis sativa L.) strains to achieve Agrobacterium-mediated transformation to enhance disease resistance. Zamir K Punja. (P-34, S23)
Meristem tips of five strains of C. sativa produced better shoot development on an initiation medium supplemented with TDZ and NAA than those supplemented with GA3, BAP or TDZ alone. Rooting media that included IBA produced 76% rooting. Callus was produced on MS with TDZ and NAA from petiole and leaf explants.
Regeneration of shoots from mature and immature inflorescences of Cannabis sativa. Gregory Golenia, K Puinno, E Boudko, C Downey, AMP Jones. (P-3029, pg 491)
Selection of candidate phenotypes of Cannabis are chosen at the late flowering stage, when ability to reproduce asexually via stem cuttings is lost. These lines are therefore cloned prior to flowering and maintained in a separate location. As an alternative method, mature and immature infloresences were cultured. These produced increasing number of shoot clusters with increasing TDZ concentration in the media. These regenerated shoots were subcultured, rooted, acclimatized and analyzed for cannibinoid and terpene content.
Cannabionoid analysis of micropropagated Cannabis sativa L. varieties using gas chromatography-flame ionization detector (GC-FID). Suman Chandra, H Lata, MM Radwan, CG Majumdar, and MA ElSohly. (P-3026, pg 490)
Cannabis has gained attention for its potential for drug abuse but also as a medicinal plant that contains valuable secondary metabolites. The plant has been reported to contain more than 500 different compounds including at least 120 cannabiunoids. GC-FID analysis of micropropagated female plants allowed categorization into high THC, high CBD and intermediate classes. This clear chemotaxonomic discrimination allows for standardization of medical-grade biomass.
Cryopreservation of in vitro Cannabis sativa L. nodal segments using vitrification. Cassandra Downey, G Golenia, E Boudko, AMP Jones. (P-1022, S38)
Due to the innate dioecious and heterozygous nature of Cannabis and unavailability of breeding programmes, the propagation of favourable genotypes commonly occurs via stem cuttings. Cryopreservation allows for long-term storage of these valuable genotypes in a relatively small space. Methods used to cryopreserve cannabis nodal cuttings to produce 70% survival and 53% regeneration (which included 10 days in the dark on regeneration) were outlined. Cryopreserved lines were grown out to flower production.
Cannabis sativa synthetic seed cold preservation and regeneration. Angelo Alvarez, N Senn. (P-2008, S53)
Encapsulation methods are being developed to maintain the large number of varieties in circulation in order to preserve the genetic diversity of Cannabis sativa. Vegetative shoot tips and node cuttings encapsulated in sodium alginate beads and stored at 4oC for 50 days survived. These synthetic seeds regenerated on in vitro media but not in sterile potting medium. This method can be used to reduce the space needed to store cannabis varieties.
Benefits of ex vitro rooting in Cannabis sativa. Norman Senn, A Alvarez. (P-2012, S55)
An ex vitro rooting method was developed for five Cannabis varieties. Rooting ex vitro (by combing stage II and III) reduced the time required to produce rooted cuttings of Cannabis.
Comparative anti-inflammatory activity of whole root, stem, leaf and flower extracts from female and male CBD hemp (Cannabis sativa L.) plants. Laura GA Esposito, E Overbaugh, J Xiong, T Rathinasabapathy, S Komarmytsky, DA Esposito. (P-2061, S72)
Industrial hemp has many applications – textiles, plastics, fuel, agricultural, nutritional and therapeutic. The distribution and accumulation of 11 major cannabinoids in CBD-rich hemp plants were quantified. The anti-inflammatory activity of hemp flowers was higher than other parts of the plant.
Phytotoxicity of streptomycin to Cannabis sativa. Savannah St Clair, N Senn. (P-3034, pg 493)
This purpose of this research was to determine the optimal concentration of streptomycin for Cannabis tissue culture in case it is needed to preserve elite lines. Streptomycin is recorded as being among the least phytotoxic antibiotics. While streptomycin was very effective as an antibacterial, leaf phytotoxic symptoms were observed and shoot growth was inhibited. The Cannabis strains tested displayed varying tolerances to the antibiotic treatment.
Organizations represented by these presentations are:
- Carleton University
- Canopy Growth Corporation
- Emergent Cannabis Sciences
- Federal University of Vicosa
- Front Range Biosciences
- Knockout Genetics, LLC
- New West Genetics
- North Carolina State University
- Simon Fraser University
- University of Guelph
- University of Mississippi
These presentations have been made possible by the following support:
- Green Roads
Next year promises to be just as exciting!
Submitted by Dr. Sylvia Mitchell, SIVB member and Senior Lecturer, Medicinal Plant Research Group, Biotechnology Centre, University of the West Indies, Mona Campus, Jamaica