Skip to main content
SpringerLink
Log in

Physical factors increased quantity and quality of micropropagated shoots of Cannabis sativa L. in a repeated harvest system with ex vitro rooting

  • Plant Tissue Culture
  • Published:
In Vitro Cellular & Developmental Biology - Plant Aims and scope Submit manuscript

Abstract

Micropropagation is a preferred method to propagate clean, clonal stock plants. Subculture is labor intensive and costly. In vitro hedging can reduce hood labor and was demonstrated with cannabis (Cannabis sativa L. ‘US Nursery Cherry 1’) using both apical and nodal explants at four different photosynthetic photon flux densities (25–167 μmol m−2 s−1) in vessels with vented or non-vented closures. The numbers of harvested shoot tips from four repeated 3-wk cutting cycles without subculture and the quality of the harvested shoot tips during ex vitro rooting in phenolic foam plugs were observed. The number of shoot tips harvested in non-vented vessels increased over four repeated cycles; however, there was a decline in number of shoot tips harvested from vented vessels in the fourth cycle due to excessive drying and collapse of the agar matric. The number of shoot tips harvested through repeated cycles was increased with light intensity. Nearly 100% of the micro-cuttings from optimal light and ventilation rooted ex vitro. Most plantlets had roots that penetrated the exterior surfaces of the plug after 2 wk. The number of leaves per rooted plantlet ex vitro increased with light intensity in vitro. Micropropagation labor efficiency could be improved by using a multi-cycle cutting process which would allow the same material to be repeatedly cut from rooted bases instead of subcultured. In a system with strong apical dominance, enhanced axillary divisions of shoot tip explants over successive cycles of cutting, without the need of exogenous PGRs, were further increased by appropriate in vitro factors, light, and ventilation, presenting labor saving potential compared to standard, single-cut system in common use.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Figure 1.
Figure 2.
Figure 3.
Figure 4.
Figure 5.
Figure 6.
Figure 7.

Similar content being viewed by others

References

  • Adelberg J, Kroggel M, Toler J (2000) Physical environment in vitro affects laboratory and nursery growth of micropropagated Hostas. HortTechnology 10:754–757

    Article  Google Scholar 

  • Ahloowalia BS, Savangikar VA (2004) Low cost options for energy and labour. In: Low cost options for tissue culture technology in developing countries. IAEA-TECDOC-1384 pp. 41–45. International Atomic Energy Agency, Vienna, Austria

  • Aitken-Christie J, Davies HE (1988) Development of a semi-automated micropropagation system. Acta Hort. 230:81–87

  • Aitken-Christie J, Jones C (1987) Towards automation: radiata pine shoot hedges in vitro. Plant Cell Tiss Org Cult 8:185–196

  • Chandra S, Lata H, El Sohly MA (Eds.). Cannabis sativa L.-botany and biotechnology In: Cannabis sativa L.—botany and biotechnology. Springer, Cham, pp. 79–100(2017)

    Google Scholar 

  • 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.). Ind Crop Prod 83:61–65

    Article  Google Scholar 

  • Debergh PC, Topoonyanont N, Van Huylenbroeck J, Moreira da Silva H, Oyaert E (2000) Preparation of microplants for ex vitro establishment. Acta Hort. 530:269–275

  • Desjardins Y (1994) Photosynthesis in vitro-on the factors regulating CO2 assimilation in micropropagation systems. Env Eff Con Plant Tiss Cult 393:45–62

  • Driver JA, Kuniyuki AH (1984) In vitro propagation of Paradox walnut rootstock. HortScience 19:507–509

    Google Scholar 

  • Economou AS (2011) From microcutting rooting to microplant establishment: key points to consider for maximum success in woody plants. Acta Hort 988:43–56

  • El-Hawaz RF, Adelberg J, Naylor-Adelberg J, Eisenreich R, Van der Meij J (2019) The effect of slow-growth strategy on a production of Petunia× hybrida Vilm. microcuttings. In Vitro Cell Dev Biol - Plant 55:433–441

  • ElSohly MA, Radwan MM, Gul W, Chandra S, Galal A (2017) Phytochemistry of Cannabis sativa L. In: Phytocannabinoids. Springer, Cham, pp 1–36

    Google Scholar 

  • Fujiwara K, Kozai T (1995) Physical microenvironment and its effects. In: Aitken-Cristie J, Kozai T, Smith MAL (eds) Automation and Environmental Control in Plant Tissue Culture. Kluwer Academic Publishers, Dordrecht, pp 319–369

    Chapter  Google Scholar 

  • Hazarika BN (2003) Acclimatization of tissue-cultured plants. Curr Sci 85:1704–1712

    CAS  Google Scholar 

  • Hoagland DR, Snyder WC (1933) Nutrition of strawberry plants under controlled conditions. AmerSoc Hort Sci 30:288–294

  • Kitaya Y, Fukuda O, Kozai T, Kirdmanee C (1995) Effects of light intensity and lighting direction on the photoautotrophic growth and morphology of potato plantlets in vitro. Sci Hortic 62:15–24

    Article  Google Scholar 

  • Kozai T, Iwanami Y (1988) Effects of CO2 enrichment and sucrose concentration under high photon fluxes on plantlet growth of carnation (Dianthus caryophyllus L.) in tissue culture during the preparation stage. J Japanese Soc Hort Sci 57:279–288

  • Kozai T, Koyama Y, Watanabe I (1988) Multiplication and rooting of potato plantlets in vitro with sugar-free medium under high photosynthetic photon flux Acta Hort 230:121–127

  • Lata H, Chandra S, Khan I, ElSohly MA (2009) Thidiazuron-induced high-frequency direct shoot organogenesis of Cannabis sativa L. In Vitro Cell Dev Biol - Plant 45:12–19

  • Lata H, Chandra S, Khan IA, ElSohly MA (2017) Micropropagation of Cannabis sativa L.—an update. In: Chandra S, Lata H, El Sohly M (eds) Cannabis sativa L. - Botany and Biotechnology. Springer, Cham, pp 285–297

    Chapter  Google Scholar 

  • Lata H, Chandra S, Techen N, Khan IA, ElSohly MA (2016) In vitro mass propagation of Cannabis sativa L.: a protocol refinement using novel aromatic cytokinin meta-topolin and the assessment of eco-physiological, biochemical and genetic fidelity of micropropagated plants. J Appl Res Med Aromat Plant 3:18–26

    Google Scholar 

  • Lee TJ, Zobayed SMA, Firmani F, Park EJ (2019) A novel automated transplanting system for plant tissue culture. Biosyst Eng 181:63–72

  • Mestinšek Mubi Š, Svetik S, Flajšman M, Murovec J (2020) In vitro tissue culture and genetic analysis of two high-CBD medical cannabis (Cannabis sativa L.) breeding lines. Genetika 52(3):925–941

    Article  Google Scholar 

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

    Article  CAS  Google Scholar 

  • Neto AR, Chagas EA, Costa BNS, Chagas PC, Vendrame WA (2020) Photomixotrophic growth response of sugarcane in vitro plantlets using different light intensities and culture vessel types. In Vitro Cell Dev Biol-Plant 56:504–514

  • Page SR, Monthony AS, Jones AMP (2020) Basal media optimization for the micropropagation and callogenesis of Cannabis sativa L. BioRxiv 1–23

  • Piunno KF, Golenia G, Boudko EA, Downey C, Jones AMP (2019) Regeneration of shoots from immature and mature inflorescences of Cannabis sativa. Can J Plant Sci 99:556–559

    Article  Google Scholar 

  • Rohr R, Iliev I, Scaltsoyinnes A, Tsoulpha P (2001) Acclimatization of micropropagated forest trees. Acta Hort 616:59–69

  • da Silva JAT, Dobránszki J, Ross S (2013) Phloroglucinol in plant tissue culture. In Vitro Cell Dev Biol - Plant 49:1–16

  • Sluis CJ (2008) Integrating automation technologies with commercial micropropagation. In: Plant Tissue Culture Engineering. Springer, Dordrecht, pp 231–251

    Google Scholar 

  • Smith M, Palta J, McCown B (1986) Comparative anatomy and physiology of microcultured, seedling, and greenhouse-grown Asian white birch. J Amer Soc Hort Sci 111:437–442

  • Van Staden J, Fennell CW, Taylor NJ (2006) Plant stress in vitro: the role of phytohormones. Acta Hort 725:55–61

  • Wang R, He LS, Xia B, Tong JF, Li N, Peng F (2009) A micropropagation system for cloning of hemp (Cannabis sativa L.) by shoot tip culture. Pak J Bot 41:603–608

    CAS  Google Scholar 

  • Wetzstein H, Sommer H (1982) Leaf anatomy of tissue cultured Liquidambar styraciflua (Hamamelidaceae) during acclimatization. Amer J Bot 69:1579–1586

  • Wróbel T, Dreger M, Wielgus K, Słomski R (2020) Modified nodal cuttings and shoot tips protocol for rapid regeneration of Cannabis sativa L. J Natl Fibers 19:1–10

    Article  Google Scholar 

  • Ziv M (1995) In vitro acclimatization. In: Aitken-Cristie J, Kozai T, Smith MAL (eds) Automation and environmental control in plant tissue culture. Kluwer Academic Publishers, Dordrecht, pp 493–516

    Chapter  Google Scholar 

  • Zobayed SMA (2005) Ventilation in micropropagation. In: Photoautotrophic (sugar-free medium) micropropagation as a new micropropagation and transplant production system. Springer, Dordrecht, pp 147–186

    Chapter  Google Scholar 

  • Zobayed SMA, Armstrong J, Armstrong W (2001) Micropropagation of potato: evaluation of closed, diffusive and forced ventilation on growth and tuberization. Ann Bot 87:53–59

    Article  CAS  Google Scholar 

Download references

Funding

This research was supported by a grant from the South Carolina Department of Agriculture.

Author information

Authors and Affiliations

  1. Department of Plant Environmental Science, Clemson University, 275 Poole Agricultural Center, Clemson, SC, 29634, USA

    Ryan Murphy & Jeffrey Adelberg

Authors
  1. Ryan Murphy
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Jeffrey Adelberg
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Ryan Murphy.

Ethics declarations

Mr. Murphy reports grants from South Carolina Department of Agriculture, during the conduct of the study; In addition, Dr. Adelberg has a patent Multicutter system for plant micropropagation pending.

Dr. Adelberg reports grants from SC Department of Agriculture, during the conduct of the study.

Additional information

Editor: Pamela Weathers

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Murphy, R., Adelberg, J. Physical factors increased quantity and quality of micropropagated shoots of Cannabis sativa L. in a repeated harvest system with ex vitro rooting. In Vitro Cell.Dev.Biol.-Plant 57, 923–931 (2021). https://doi.org/10.1007/s11627-021-10166-4

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s11627-021-10166-4

Keywords

Search

Navigation