Journal of Medicinal and Aromatic Plant Sciences

Volume: 45 Issue: 1

  • subscription
  • Review Article

Exploring methodologies in Cannabis tissue-culture and genetic transformation: Opportunities and obstacles

SHUKLA, D.*

*Tissue-culture and Transformation Facility, Plant Biotechnology Division, CSIR – Central Institute of Medicinal and Aromatic Plants, Picnic Spot road Kukrail, Lucknow – 226015, Uttar Pradesh, India

Email: [email protected]

Year: 2023, Page: 1-16, Doi: https://doi.org/10.62029/jmaps.v45i1.shukla

Received: May 12, 2023 Accepted: June 17, 2023 Published: Oct. 27, 2023

Abstract

In recent years, the growing interest in Cannabis sativa L., particularly its medicinal and aromatic properties, has propelled advancements in its tissue culture and genetic transformation techniques. This review delineates the significant strides and persistent challenges in the field, offering a comprehensive overview of the current methodologies and their implications. It discusses the synergistic effects of Thidiazuron (TDZ) and Naphthaleneacetic acid (NAA) in the Murashige and Skoog (MS) medium as well as the use of meta-Topolin (mT). This synthetic cytokinin (mT) facilitates a high induction frequency and many shoots per explant. It introduces a time-efficient and resource-optimized pathway for Cannabis micropropagation and germplasm conservation. The genetic transformation in Cannabis was predominantly facilitated through Agrobacterium-mediated transformation, a cornerstone technique that enabled the integration of foreign genes into the plant genome. Regulatory implications associated with gene editing in Cannabis sativa are highlighted. Despite these advancements, the field grapples with several challenges, including the recalcitrant nature of Cannabis, especially regarding in vitro propagation or genetic transformation, the genotypic specificity of regeneration protocols, and the reproducibility of existing methods. The complexity of the Cannabis genome, characterized by a high degree of polymorphism and multiple copies of specific genes, further exacerbates these challenges. Moreover, the current research landscape is marred by a lack of standardized protocols and variable responses among different Cannabis varieties, necessitating more robust and universally applicable protocols. This review underscores the pressing need for further research to optimize protocols for higher efficiency and to develop suitable systems for in-vitro plantlet regeneration.

Keywords: Cannabis sativa, Genetic transformation, Hemp, Regeneration, Tissue culture

References

Abel, E. L. (1980). Marihuana: The first twelve thousand years. New York, NY: Plenum Press.

Adhikary, D., Kulkarni, M., El-Mezawy, A., Mobini, S., Elhiti, M., Gjuric, R., Ray, A., Polowick, P., Slaski, J. J., Jones, M. P., & Bhowmik, P. (2021). Medical cannabis and industrial hemp tissue culture: Present status and future potential. Frontiers in Plant Science, 12, 627240. https://doi.org/10.3389/fpls.2021.627240

Adinoff, B., & Reiman, A., (2019). Implementing social justice in the transition from illicit to legal Cannabis. American Journal of Drug and Alcohol Abuse, 45, 673–688.

Chand, S., & Singh, A. K. (2004). Plant regeneration from encapsulated nodal segments of Dalbergia sissoo Roxb., A timber-yielding leguminous tree species. Journal of Plant Physiology, 161, 237–243.

Chandra, S., Lata, H., Khan, I. A., & El Sohly, M. A. (2011). Photosynthetic response of Cannabis sativa L. to elevated levels of CO2. Physiology and Molecular Biology of Plants, 17, 291–295.

Chaohua, C., Gonggu, Z., Lining, Z., Chunsheng, G., Qing, T., Jianhua, C., Xinbo, G., Dingxiang, P., & Jianguang, S. (2016). A rapid shoot regeneration protocol from the cotyledons of hemp (Cannabis sativa L.).Industrial Crops and Products, 83, 61–65.

Chopra, I. C., & Chopra, R. N. (1957). The use of cannabis drugs in India. Bulletin of Narcotics, 9, 4–29.

Clarke, R. C., & Merlin, M. D. (2016). Cannabis domestication, breeding history, present-day genetic diversity, and future prospects. Critical Reviews in Plant Sciences, 35, 293–327.

Deguchi, M., Bogush, D., Weeden, H., Spuhler, Z., Potlakayala, S., Kondo, T., Zhang, Z. J., & Rudrabhatla, S. (2020). Establishment and optimization of a Hemp (Cannabis sativa L.) agroinfiltration system for gene expression and silencing studies. Scientific Reports, 10, 3504.

ElSohly, M. A., Radwan, M. M., Gul, W., Chandra, S., & Galal, A. (2017). Phytochemistry of Cannabis sativa L. In A. D. Kinghorn, H. Falk, S. Gibbons, J. Kobayashi (Eds.), Phytocannabinoids, (pp. 1–36). Springer International Publishing.

Feeney, M., & Punja, Z. K. (2003). Tissue culture and Agrobacterium-mediated transformation of hemp (Cannabis sativa L.). In Vitro Cellular & Developmental Biology – Plant, 39, 578–585.

Galán-Ávila, A., García-Fortea, E., Prohens, J., & Herraiz, F. J. (2020). Development of a direct in vitro plant regeneration protocol from Cannabis sativa L. seedling explants: Developmental morphology of shoot regeneration and ploidy level of regenerated plants. Frontiers in Plant Science, 11, 645.

Galán-Ávila, A., Gramazio, P., Ron, M., Prohens, J., & Herraiz, F. J. (2021). A novel and rapid method for Agrobacterium-mediated production of stably transformed Cannabis sativa L. plants. Industrial Crops and Products, 170, 113691.

Hesami, M., Baiton, A., Alizadeh, M., Pepe, M., Torkamaneh, D., & Jones, A. M. P. (2021). Advances and perspectives in tissue culture and genetic engineering of Cannabis. International Journal of Molecular Sciences, 22, 5671.

Ioannidis, K., Tomprou, I., & Mitsis, V. (2022). An alternative in vitro propagation protocol of Cannabis sativa L. (Cannabaceae) presenting efficient rooting for commercial production. Plants, 11, 1333.

Iversen, L. L. (2008). The science of marijuana. Oxford University Press.

Kuddus, M., Ginawi, I., & AlHazimi, A. (2013). Cannabis sativa: An ancient wild edible plant of India. Emirates Journal of Food and Agriculture, 25, 736.

Lata, H., Chandra, S., Khan, I. A., & ElSohly, M. A. (2010). High frequency plant regeneration from leaf-derived callus of high Δ9-tetrahydrocannabinol yielding Cannabis sativa L. Planta Medica, 76, 1629–1633.

Lata, H., Chandra, S., Khan, I. A., & ElSohly, M. A. (2009). Propagation through alginate encapsulation of axillary buds of Cannabis sativa L.—an important medicinal plant. Physiology and Molecular Biology of Plants, 15, 79–86.

Lata, H., Chandra, S., Khan, I. A., & ElSohly, M. A. (2009). Thidiazuron-induced high-frequency direct shoot organogenesis of Cannabis sativa L. In Vitro Cellular & Developmental Biology – Plant, 45, 12–19.

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 S. M. Jain (Ed.), Protocols for in vitro cultures and secondary metabolite analysis of aromatic and medicinal plants (pp. 275–288). Springer New York.

Lubell-Brand, J. D., Kurtz, L. E., & Brand, M. H. (2021). An in vitro–ex vitro micropropagation system for hemp (Cannabis sativa L.). HortTechnology, 31, 199–207.

Monthony, A. S., Kyne, S. T., Grainger, C. M., & Jones, A. M. P. (2021). Recalcitrance of Cannabis sativa to de novo regeneration: A multi-genotype replication study. PLOS ONE, 16, e0235525.

Monthony, A. S., Page, S. R., Hesami, M., & Jones, A. M. P. (2021). The past, present and future of Cannabis sativa tissue culture. Plants, 10, 185.

Movahedi, M., Ghasemi-Omran, V., & Torabi, S. (2015). The effect of different concentrations of TDZ and BA on in vitro regeneration of Iranian Cannabis sativa using cotyledon and epicotyl explants. Journal of Plant Molecular Breeding, 3.

Musio, S., Müssig, J., & Amaducci, S. (2018). Optimizing hemp fiber production for high performance composite applications. Frontiers in Plant Science, 9, 1702.

Page, S. R. G., Monthony, A. S., & Jones, A. M. P. (2020). DKW basal salts improve micropropagation and callogenesis compared to MS basal salts in multiple commercial cultivars of Cannabis sativa. Plant Biology (preprint).

Piunno, K. F., Golenia, G., Boudko, E. A., Downey, C., & Jones, A. M. P. (2019). Regeneration of shoots from immature and mature inflorescences of Cannabis sativa. Canadian Journal of Plant Science, 99, 556–559.

Raharjo, T. J., Eucharia, O., Chang, W.-T., & Verpoorte, R. (2010). Callus induction and phytochemical characterization of Cannabis sativa cell suspension cultures. Indonesian Journal of Chemistry, 6, 70–74.

Rai, M. K., Jaiswal, V. S., & Jaiswal, U. (2008). Encapsulation of shoot tips of Psidium guajava L. for short-term storage and germplasm exchange. Scientia Horticulturae, 118, 33–38.

Rihan, H., Kareem, F., El-Mahrouk, M., & Fuller, M. (2017). Artificial seeds (principle, aspects and applications). Agronomy, 7, 71.

Slusarkiewicz-Jarzina, 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 Biologica Cracoviensia Series Botanica, 47, 145–151.

Smýkalová, I., Vrbová, M., Cvečková, M., Plačková, L., Žukauskaitė, A., Zatloukal, M., Hrdlička, J., Plíhalová, L., Doležal, K., & Griga, M. (2019). The effects of novel synthetic cytokinin derivatives and endogenous cytokinins on the in vitro growth responses of hemp (Cannabis sativa L.) explants. Plant Cell, Tissue and Organ Culture, 139, 381–394.

Wawrosch, C., & Zotchev, S. B. (2021). Production of bioactive plant secondary metabolites through in vitro technologies—Status and outlook. Applied Microbiology and Biotechnology, 105, 6649–6668.

Zhang, X., Xu, G., Cheng, C., Lei, L., Sun, J., Xu, Y., Deng, C., Dai, Z., Yang, Z., Chen, X., Liu, C., Tang, Q., & Su, J. (2021). Establishment of an Agrobacterium-mediated genetic transformation and CRISPR/Cas9-mediated targeted mutagenesis in hemp (Cannabis sativa L.). Plant Biotechnology Journal, 19, 1979–1987.

 

Cite this article

Shukla, D. 2023. Exploring methodologies in Cannabis tissue-culture and genetic transformation: Opportunities and obstacles. Journal of Medicinal and Aromatic Plant Sciences, 45(1), 1–16.

Views
3349
Downloads
7
Citations