Advertisement

Plant Cell, Tissue and Organ Culture (PCTOC)

, Volume 125, Issue 1, pp 177–181 | Cite as

Reducing etiolation-like effect and flowering in an in vitro micropropagated Trifolium resupinatum elite genotype

  • Olívia Campos Costa
  • Daniela Lopes Oliveira
  • Ana Rita Silva
  • Ana Barradas
  • João Paulo Crespo
  • Ana Sofia DuqueEmail author
  • Pedro Fevereiro
Research Note

Abstract

There is an ever-increasing demand for high quality forage legumes, essential to feed livestock. Micropropagation may be useful to preserve elite allelic compositions in allogamous and auto-incompatible forage legume species, as is the case of Trifolium resupinatum L. (Persian clover). Etiolation-like phenotypes in in vitro cultured plants, with long weak stems and sparse leaves, together with flowering, compromises the long term establishment and maintenance of explants and the adequate development of plants when transferred to ex vitro. Aiming to develop a solution to limit the impact of in vitro etiolation-like effects, flowering and plant viability, we used a very prone to etiolate elite T. resupinatum genotype maintained through in vitro propagation for over 1 year. Stem segments were subcultured in Murashige and Skoog (MS) supplemented with 2.22, 4.56 or 9.12 µM of zeatin; with 4.44 or 8.88 µM of benzyladenine or with 2.89 or 14.44 µM of gibberellic acid. Gibberellic acid supplementation was detrimental and ultimately led to the death of the explants. Supplementation with 2.22 µM zeatin produce no significant differences from control; however higher concentrations of zeatin, as well as of benzyladenine, diminished the longer internodes and increased the development of shoots with more stout stems. Although rooting was prevented by cytokinins, it was successfully induced when explants were transferred to MS media without plant growth regulators. The 9.12 µM zeatin condition was the most favorable since it reversed the etiolation-like phenotype, inhibited in vitro flowering and improved acclimation from 28.7 to 90 % in the FTTr07.13 T. resupinatum genotype.

Keywords

Etiolation-like effect In vitro flowering Trifolium resupinatum Elite genotype 

Notes

Acknowledgments

Authors acknowledge financial support from Fundação para a Ciência e a Tecnologia (Lisboa, Portugal) through the research Grant SFRH/BPD/74784/2010 (Duque AS), the Research unit GREEN-it “Bioresources for Sustainability” (UID/Multi/04551/2013) and PRODER (4.1) Project Micropropelite (No. 020536053572). We acknowledge Dr. Susana Araújo for revising the manuscript.

References

  1. Abberton MT (2007) Interspecific hybridization in the genus Trifolium. Plant Breed 126:337–342CrossRefGoogle Scholar
  2. Araújo SS, Duque AS, Santos DM, Fevereiro P (2004) An efficient transformation method to regenerate a high number of transgenic plants using a new embryogenic line of Medicago truncatula cv. Jemalong. Plant Cell, Tissue Organ Cult 78:123–131CrossRefGoogle Scholar
  3. Araújo SS, Beebe S, Crespi M, Delbreil B, González EM, Gruber V, Lejeune-Henaut I, Link W, Monteros MJ, Prats E, Rao I, Vadez V, Vaz Patto MC (2015) Abiotic stress responses in legumes: strategies used to cope with environmental challenges. Crit Rev Plant Sci 34(1–3):237–280CrossRefGoogle Scholar
  4. Aremu AO, Bairu MW, Dolezˇal K, Finnie JF, Van Staden J (2012) Topolins: A panacea to plant tissue culture challenges? Plant Cell, Tissue Organ Cult 108:1–16CrossRefGoogle Scholar
  5. Aremu AO, Plačková L, Bairu MW, Novák O, Plíhalová L, Doležal K, Finnie JF, Van Staden J (2014) How does exogenously applied cytokinin type affect growth and endogenous cytokinins in micropropagated Merwilla plumbea? Plant Cell, Tissue Organ Cult 118:245–256CrossRefGoogle Scholar
  6. Chory J, Reinecke D, Sim S, Washburn T, Brenner M (1994) A role for cytokinins in de-etiolation in Arabidopsis (det mutants have an altered response to cytokinins). Plant Physiol 104:339–347PubMedPubMedCentralGoogle Scholar
  7. Dill A, Sun T (2001) Synergistic derepression of gibberellin signaling by removing RGA and GAI function in Arabidopsis thaliana. Genetics 159:777–785PubMedPubMedCentralGoogle Scholar
  8. Duque AS, Barradas A, Godinho B, Silva AR, Araújo SS, Crespo JP, Fevereiro P (2015) Development of protocols for micropropagation of elite genotype forage allogamous legume species. Acta Hortic 1083:409–413CrossRefGoogle Scholar
  9. Fleet CM, Sun T (2005) A DELLAcate balance: the role of gibberellin in plant morphogenesis. Curr Opin Plant Biol 8:77–85CrossRefPubMedGoogle Scholar
  10. Gaspar T, Kevers C, Penel C, Greppin H, Reid DM, Thorpe TA (1996) Plant hormones and plant growth regulators in plant tissue culture. Vitr Cell Dev Biol Plant 32:272–289CrossRefGoogle Scholar
  11. George EF, Hall MA, De Klerk G-J (eds) (2008) Plant tissue culture procedure - background. In: Plant propagation by tissue culture, vol 1, 3rd edn. Springer, The Netherlands, pp 1–28Google Scholar
  12. Goldberg-Moeller R, Shalom L, Shlizerman L, Samuels S, Zur N, Ophir R, Blumwald E, Sadka A (2013) Effects of gibberellin treatment during flowering induction period on global gene expression and the transcription of flowering-control genes in Citrus buds. Plant Sci 198:46–57CrossRefPubMedGoogle Scholar
  13. Greenboim-Wainberg Y, Maymon I, Borochov R, Alvarez J, Olszewski N, Ori N, Eshed Y, Weiss D (2005) Cross talk between gibberellin and cytokinin: the Arabidopsis GA response inhibitor SPINDLY plays a positive role in cytokinin signaling. Plant Cell 17:92–102CrossRefPubMedPubMedCentralGoogle Scholar
  14. Kurepin LV, Pharis RP (2014) Light signaling and the phytohormonal regulation of shoot growth. Plant Sci 229:280–289CrossRefPubMedGoogle Scholar
  15. Loberant B, Altman A (2010) Micropropagation of plants. In: Flickinger MC (ed) Encyclopedia of industrial biotechnology: bioprocess, bioseparation, and cell technology. Wiley, New York, pp 1–17Google Scholar
  16. Mobini SH, Lulsdorf M, Warkentin TD, Vandenberg A (2015) Plant growth regulators improve in vitro flowering and rapid generation advancement in lentil and faba bean. Vitr Cell Dev Biol Plant 51:71–79CrossRefGoogle Scholar
  17. Murashige T, Skoog F (1962) A revised medium for rapid growth and bio assays with tobacco tissue cultures. Physiol Plant 15:473–497CrossRefGoogle Scholar
  18. Mutasa-Gottgens E, Hedden P (2009) Gibberellin as a factor in floral regulatory networks. J Exp Bot 60:1979–1989CrossRefPubMedGoogle Scholar
  19. Ribalta FM, Croser JS, Erskine W, Finnegan PM, Lulsdorf MM, Ochatt SJ (2014) Antigibberellin-induced reduction of internode length favors in vitro flowering and seed-set in different pea genotypes. Biol Plant 58(1):39–46CrossRefGoogle Scholar
  20. Symons GM, Reid JB (2003) Interactions between light and plant hormones during de-etiolation. J Plant Growth Regul 22:3–14CrossRefGoogle Scholar
  21. Thorpe TA (2007) History of plant tissue culture. Mol Biotechnol 37(2):169–180CrossRefPubMedGoogle Scholar
  22. Weiss D, Ori N (2007) Mechanisms of cross talk between gibberellin and other hormones. Plant Physiol 144:1240–1246CrossRefPubMedPubMedCentralGoogle Scholar

Copyright information

© Springer Science+Business Media Dordrecht 2015

Authors and Affiliations

  • Olívia Campos Costa
    • 1
    • 3
  • Daniela Lopes Oliveira
    • 1
  • Ana Rita Silva
    • 3
  • Ana Barradas
    • 3
  • João Paulo Crespo
    • 3
  • Ana Sofia Duque
    • 1
    Email author
  • Pedro Fevereiro
    • 1
    • 2
  1. 1.Plant Cell Biotechnology Laboratory, Green-it UnitInstituto de Tecnologia Química e Biológica António Xavier (ITQB)OeirasPortugal
  2. 2.Departamento de Biologia VegetalFaculdade de Ciências da Universidade de LisboaLisbonPortugal
  3. 3.Fertiprado - Sementes e Nutrientes Lda.VaiamontePortugal

Personalised recommendations