Skip to main content
SpringerLink
Log in

An efficient and reproducible Agrobacterium-mediated genetic transformation method for the ornamental monocotyledonous plant Ornithogalum dubium Houtt

  • Original paper
  • Published:
Plant Growth Regulation Aims and scope Submit manuscript

Abstract

Ornithogalum is a genus from the Hyacinthaceae (Asparagaceae) family that comprises about 200 species with remarkable white, yellow, or orange flowers that display exceptional vase life. These properties have made it a popular cut flower and pot plant. Forward genetics approaches may be advantageous to generate novel phenotypes, but the Agrobacterium-mediated transformation of plants from this genus remains challenging. Here, a stable and efficient Agrobacterium-mediated transformation system was established for O. dubium. We found that the timing of transformation with respect to light exposure of the tissue affected transformation rates more than other tested parameters. In the transgenic plants obtained, T-DNA integrations were confirmed by polymerase chain reactions and positive plants were established in the greenhouse and displayed weak transgene expression. This study exposed an efficient platform for gene function research and germplasm improvement in O. dubium plants. The present protocol is now available for the development of novel improved O. dubium varieties.

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

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6
Fig. 7
Fig. 8

Similar content being viewed by others

Data availability

The data underlying this article are available in the article and in its online supplemental material.

References

  • Ascough GD, Erwin JE, van Staden J (2009) Micropropagation of iridaceae—a review. Plant Cell Tissue Organ Cult 97:1–19

    Article  Google Scholar 

  • Baulcombe DC (1996) RNA as a target and an initiator of post-transcriptional gene silencing in transgenic plants. Plant Mol Biol 32:79–88

    Article  CAS  PubMed  Google Scholar 

  • Cohen A, Meredith CP (1992) Agrobacterium—mediated transformation of lilium. International Society for Horticultural Science (ISHS), Leuven, Belgium

  • Cohen A, Lipsky A, Arazi T et al (2004) Efficient genetic transformation of Lilium longiflorum and Ornithogalum dubium by particle acceleration followed by prolonged selection in liquid medium. International Society for Horticultural Science (ISHS), Leuven, Belgium

  • Cohen A, Lipsky A, Arazi T et al (2005) Particle bombardment-mediated transformation of ornithogalum dubium for ornithogalum mosaic virus resistance. In: Acta horticulturae. international society for horticultural science, pp 183–190

  • Doyle J (1991) DNA protocols for plants. In: Hewitt GM, Johnston AWB, Young JPW (eds) Molecular techniques in taxonomy. Springer, pp 283–293

    Chapter  Google Scholar 

  • Finer JJ, Vain P, Jones MW, McMullen MD (1992) Development of the particle inflow gun for DNA delivery to plant cells. Plant Cell Rep 11:323–328

    Article  CAS  PubMed  Google Scholar 

  • Garcia R, Cidade D, Castellar A et al (2007) In vitro morphogenesis patterns from shoot apices of sugar cane are determined by light and type of growth regulator. Plant Cell Tissue Organ Cult 90:181–190

    Article  Google Scholar 

  • Griesbach RJ, Meyer F, Koopowitz H (1993) Creation of new flower colors in ornithogalum via interspecific hybridization. J Am Soc Hortic Sci 118:409–414

    Article  Google Scholar 

  • Höfgen R, Willmitzer L (1988) Storage of competent cells for Agrobacterium transformation. Nucl Acids Res 16:9877

    Article  PubMed  PubMed Central  Google Scholar 

  • Hofmann NR (2016) A breakthrough in monocot transformation methods. Plant Cell 28:1989

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Hooykaas-Van Slogteren GMS, Hooykaas PJJ, Schilperoort RA (1984) Expression of Ti plasmid genes in monocotyledonous plants infected with Agrobacterium tumefaciens. Nature 311:763–764

    Article  CAS  Google Scholar 

  • Hoshi Y, Kondo M, Mori S et al (2004) Production of transgenic lily plants by Agrobacterium-mediated transformation. Plant Cell Rep 22:359–364

    Article  CAS  PubMed  Google Scholar 

  • Hussey G (1976) Plantlet regeneration from callus and parent tissue in Ornithogalum thyrsoides. J Exp Bot 27:375–382

    Article  CAS  Google Scholar 

  • Hussey G (1977) In vitro propagation of some members of the liliaceae, iridaceae and amaryllidaceae. International Society for Horticultural Science (ISHS), Leuven, Belgium

  • Hvoslef-Eide AK, Preil W (2005) Liquid culture systems for in vitro plant propagation. Springer, London

  • Kamo K (1997) Factors affecting Agrobacterium tumefaciens-mediatedgus A expression and opine synthesis in Gladiolus. Plant Cell Rep 16:389–392

    CAS  PubMed  Google Scholar 

  • Koetle MJ, Finnie JF, Balázs E, Van Staden J (2015) A review on factors affecting the Agrobacterium-mediated genetic transformation in ornamental monocotyledonous geophytes. S Afr J Bot 98:37–44

    Article  CAS  Google Scholar 

  • Kumar M, Ayzenshtat D, Marko A, Bocobza S (2021) Optimization of T-DNA configuration with UBIQUITIN10 promoters and tRNA-sgRNA complexes promotes highly efficient genome editing in allotetraploid tobacco. Plant Cell Rep. https://doi.org/10.1007/s00299-021-02796-0

    Article  PubMed  Google Scholar 

  • Kumar M, Tripathi PK, Ayzenshtat D et al (2022) Increased rates of gene-editing events using a simplified RNAi configuration designed to reduce gene silencing. Plant Cell Rep. https://doi.org/10.1007/s00299-022-02903-9

    Article  PubMed  PubMed Central  Google Scholar 

  • Langeveld SA, Gerrits MM, Derks AFLM et al (1995) Transformation of lily by Agrobacterium. Euphytica 85:97–100

    Article  CAS  Google Scholar 

  • Lipsky A, Cohen A, Ion A, Yedidia I (2014) Genetic transformation of Ornithogalum via particle bombardment and generation of Pectobacterium carotovorum-resistant plants. Plant Sci 228:150–158

    Article  CAS  PubMed  Google Scholar 

  • Lipsky A, Joshi JR, Carmi N, Yedidia I (2016) Expression levels of antimicrobial peptide tachyplesin I in transgenic Ornithogalum lines affect the resistance to Pectobacterium infection. J Biotechnol 238:22–29

    Article  CAS  PubMed  Google Scholar 

  • Littlejohn GM, Blomerus LM (1997) Evaluation of Ornithogalum genebank accessions for some characteristics of importance for breeding cut flowers or pot plants. Genet Resour Crop Evol 44:227–234

    Article  Google Scholar 

  • Luria G, Watad AA, Cohen-Zhedek Y, Borochov A (2002) Growth and flowering of ornithogalum dubium. International Society for Horticultural Science (ISHS), Leuven, Belgium

  • Mori S, Adachi Y, Horimoto S et al (2005) Callus formation and plant regeneration in various Lilium species and cultivars. In Vitro Cell Dev Biol Plant 41:783–788

    Article  Google Scholar 

  • Naik P, Nayak S (2005) Different modes of plant regeneration and factors affecting in vitro bulblet production in Ornithogalum virens. Sci Asia 31:409–414

  • Niederwieser JG, Bornman CH (2004) Role of biotechnology in the development and production of Lachenalia and Ornithogalum cultivars in South Africa. S Afr J Bot 70:47–51

    Article  Google Scholar 

  • Niederwieser JG, van de Venter HA, Robbertse PJ (1990) Embryo rescue in ornithogalum. Hort Sci 25:565–566

    Google Scholar 

  • Petti C (2020) Phloroglucinol mediated plant regeneration of Ornithogalum dubium as the sole “hormone-like supplement” in plant tissue culture long-term experiments. Plants. https://doi.org/10.3390/plants9080929

  • Pola S, Saradamani N, Ramana T (2007) Enhanced shoot regeneration in tissue culture studies of Sorghum bicolor. http://ijat-aatsea.com/pdf/Nov_V3_no2_07/11-IJAT2007_20-P%20275-286.pdf. Accessed 2 Feb 2023

  • Sarrion-Perdigones A, Vazquez-Vilar M, Palací J et al (2013) GoldenBraid 2.0: a comprehensive DNA assembly framework for plant synthetic biology. Plant Physiol 162:1618–1631

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Silhavy D, Molnár A, Lucioli A et al (2002) A viral protein suppresses RNA silencing and binds silencing-generated, 21- to 25-nucleotide double-stranded RNAs. EMBO J 21:3070–3080

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Singh V, Sergeeva L, Ligterink W et al (2019) Gibberellin promotes sweetpotato root vascular lignification and reduces storage-root formation. Front Plant Sci 10:1320

    Article  PubMed  PubMed Central  Google Scholar 

  • Sood P, Bhattacharya A, Sood A (2011) Problems and possibilities of monocot transformation. Biol Plant 55:1–15

    Article  CAS  Google Scholar 

  • Suzuki S, Supaibulwatana K, Mii M, Nakano M (2001) Production of transgenic plants of the Liliaceous ornamental plant Agapanthus praecox ssp. orientalis (Leighton) Leighton via Agrobacterium-mediated transformation of embryogenic calli. Plant Sci 161:89–97

    Article  CAS  Google Scholar 

  • Talbot MJ, White RG (2013) Methanol fixation of plant tissue for scanning electron microscopy improves preservation of tissue morphology and dimensions. Plant Methods 9:36

    Article  PubMed  PubMed Central  Google Scholar 

  • Tun OM, Lipsky A, Luzzatto Knaan T et al (2013) The plant activator BTH promotes Ornithogalum dubium and O. thyrsoides differentiation and regeneration in vitro. Biol Plant 57:41–48

    Article  CAS  Google Scholar 

  • van Emmenes L, Veale A, Cohen A, Arazi T (2008) Agrobacterium-mediated transformation of the bulbous flower ornithogalum. International Society for Horticultural Science (ISHS), Leuven, Belgium

  • Ziv M, Lilien-Kipnis H (1997) Bud cluster proliferation in bioreactor cultures of ornithogalum dubium. International Society for Horticultural Science (ISHS), Leuven, Belgium

Download references

Funding

This study was financially supported by the Chief Scientist—Ministry of Agriculture and Rural Development No. 20-01-0241.

Author information

Authors and Affiliations

  1. Institute of Plant Sciences, ARO Volcani Center, P.O. Box 15159, 7528809, Rishon LeZion, Israel

    Pankaj Kumar Tripathi, Dana Ayzenshtat, Manoj Kumar, Hanita Zemach, Iris Yedidia & Samuel E. Bocobza

Authors
  1. Pankaj Kumar Tripathi
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Dana Ayzenshtat
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Manoj Kumar
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Hanita Zemach
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Iris Yedidia
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Samuel E. Bocobza
    View author publications

    You can also search for this author in PubMed Google Scholar

Contributions

SB conceived and designed the research. PKT, DA, and MK conducted the experiments. Histological and SEM assays were performed with the assistance of HZ. SB and IY supervised the overall work and critically analyzed all the results. PKT, DA, IY, and SB wrote the manuscript. All the authors read and approved the final manuscript.

Corresponding author

Correspondence to Samuel E. Bocobza.

Ethics declarations

Conflict of interest

The authors declare they have no conflict of interest.

Additional information

Communicated by Ben Zhang.

Publisher's Note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Supplementary Information

Below is the link to the electronic supplementary material.

Supplemental Table S1

: Media used for O. dubium explant transformation. Supplemental Table S2: Hormonal combinations used for rooting in O. dubium transformed tissues. Supplemental Table S3: List of oligonucleotides used in this study. Supplemental Table S4: Effects of kanamycin concentrations on the survival of O. dubium calli. Supplemental Fig. S1: Schematic representation of the constructs used in this assay, consisting of the CaMV-35S and different UBIQUITIN10 promoters and terminators to control the NPT2 and RFP genes (Kumar et al. 2021). Supplemental Fig. S2: RFP fluorescence detected in the plants. Supplemental Fig. S3: Transformation of O. dubium using Agrobacterium at different OD600. Supplemental Fig. S4: Transformation of O. dubium using two different Agrobacterium strains (EHA105 and AGL1). Supplemental Fig. S5: RFP fluorescence detected in the plants. (DOCX 3399 kb)

Rights and permissions

Springer Nature or its licensor (e.g. a society or other partner) holds exclusive rights to this article under a publishing agreement with the author(s) or other rightsholder(s); author self-archiving of the accepted manuscript version of this article is solely governed by the terms of such publishing agreement and applicable law.

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Tripathi, P.K., Ayzenshtat, D., Kumar, M. et al. An efficient and reproducible Agrobacterium-mediated genetic transformation method for the ornamental monocotyledonous plant Ornithogalum dubium Houtt. Plant Growth Regul 101, 201–214 (2023). https://doi.org/10.1007/s10725-023-01013-0

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s10725-023-01013-0

Keywords

Search

Navigation