Skip to main content

Effects of Cytokinins on In Vitro Seed Germination and Early Seedling Morphogenesis in Lotus corniculatus L.

Journal of Plant Growth Regulation volume 25, Article number: 187 (2006) Cite this article

Abstract

We determined the effects of zeatin (ZEA), isopentenyl adenine (2iP), kinetin (KIN), benzyladenine (BA), and thidiazuron (TDZ) on seed germination, elongation of seedling shoots and roots, frequency of regeneration, and the number of regenerants per seedling in Lotus corniculatus L. Sterilized seeds were cultured in vitro on Murashige and Skoog (1962) medium containing 3% sucrose, 0.7% agar, and various cytokinins (0, 0.08, 0.22, 0.35, 0.80, 2.20, and 3.50 μM). After 30 days, seedlings were transferred to cytokinin-free medium for another 60 days. All cytokinins stimulated the rate and percentage of seed germination at least twofold in optimum concentrations; TDZ and ZEA were the most active, followed closely by BA, whereas KIN and 2iP stimulated germination in higher concentrations only. Elongation of shoots and roots was strongly inhibited at the lowest TDZ and BA concentrations, whereas ZEA, KIN, and 2iP exerted moderate, dose-dependent inhibition. The frequency of regenerant-producing seeds was highest on ZEA and BA, whereas the greatest number of regenerants per seedling was found on TDZ. It is concluded that the culture of seeds on cytokinin-containing media, followed by transfer to cytokinin-free medium, is a suitable procedure for rapid production of a large number of uniform regenerants. The presumed role of particular cytokinins is discussed.

This is a preview of subscription content, access via your institution.

Access options

Buy single article

Instant access to the full article PDF.

USD 39.95

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

Figure 1.
Figure 2.
Figure 3.
Figure 4.

References

  • Arcioni S, Mariotti D, Damiani F, Pezzotti M. 1988. Bird’s foot trefoil (Lotus corniculatus L), crownvetch (Coronilla varia L) and sainfoin (Onobrychis viciifolia Scop). In YPS Bajaj (ed), Biotechnology in Agriculture and Forestry, Vol. 6, Crops II, Springer-Verlag, Berlin Heidelberg, p 548–572

    Google Scholar 

  • Atici Ö, Ağar G, Battal P. 2005. Changes in phytohormone contents in chickpea seeds germinating under lead or zink stress. Biol Plant 49:215–222

    CAS  Article  Google Scholar 

  • Auer CA, Motyka V, Březinová A, Kaminek M. 1999. Endogenous cytokinin accumulation and cytokinin oxidase activity during shoot organogenesis of Petunia hybrida. Physiol Plant 105:141–147

    CAS  Article  Google Scholar 

  • Badzian T, Rybczyński JJ. 1994. Cytokinin control of shoot regeneration in root segment culture of Lotus corniculatus seedling. Acta Physiol Plant 16:61–67

    CAS  Google Scholar 

  • Bilyeu KD, Laskey JG, Morris RO. 2003. Dynamics of expression and distribution of cytokinin oxidase/dehydrogenase in developing maize kernels. Plant Growth Regul 39:195–203

    CAS  Article  Google Scholar 

  • Chaitanya KSK, Naithani SC. 1998. Kinetin-mediated prolongation of viability in recalcitrant sal (Shorea robusta Gaertn f.) seeds at low temperature: role of kinetin in delaying membrane deterioration during desiccation-induced injury. J Plant Growth Regul 17:63–69

    CAS  Article  Google Scholar 

  • Chiwocha SDS, Cutler AJ, Abrams SR, Ambrose SJ, Yang J, et al. 2005. The etr1-2 mutation in Arabidopsis thaliana affects the abscisic acid, auxin, cytokinin and gibberellin metabolic pathways during maintenance of seed dormancy, moist-chilling and germination. Plant J 42:35–48

    PubMed  CAS  Article  Google Scholar 

  • Gadallah MAA, El-Enany AE. 1999. Role of kinetin in alleviation of copper and zinc toxicity in Lupinus termis plants. Plant Growth Regul 29:151–160

    CAS  Article  Google Scholar 

  • Galuszka P, Frébort I, Šebela M, Peč P. 2000. Degradation of cytokinins by cytokinin oxidases in plants. Plant Growth Regul 32:315–327

    CAS  Article  Google Scholar 

  • Galuszka P, Frébort I, Šebela M, Sauer P, Jacobsen S, et al. 2001. Cytokinin oxidase or dehydrogenase? Mechanism of cytokinin degradation in cereals. Eur J Biochem 268:450–461

    PubMed  CAS  Article  Google Scholar 

  • Gaudinová A, Dobrev PI, Šolcová B, Novák O, Strnad M, et al. 2005. The involvement of cytokinin oxidase/dehydrogenase and zeatin reductase in regulation of cytokinin levels in pea (Pisum sativum L.) leaves. J Plant Growth Regul 24:188–200

    Article  CAS  Google Scholar 

  • Gerhäuser D, Bopp M. 1990. Cytokinin oxidases in mosses. 2. Metabolism of kinetin and benzyladenine in vitro. J Plant Physiol 135:714–718

    Google Scholar 

  • Gulzar S, Khan MA. 2002. Alleviation of salinity-induced dormancy in perennial grasses. Biol Plant 45:617–619

    CAS  Article  Google Scholar 

  • Hare PD, van Staden J. 1994. Cytokinin oxidase: Biochemical features and physiological significance. Physiol Plant 91:128–136

    CAS  Article  Google Scholar 

  • Khan MA, Ungar IA. 1997. Alleviation of seed dormancy in the desert forb Zygophyllum simplex L. from Pakistan. Ann Bot 80:395–400

    CAS  Article  Google Scholar 

  • Khan MA, Gul B, Weber DJ. 2004. Action of plant growth regulators and salinity on seed germination of Ceratoides lanata. Can J Bot 82:37–42

    CAS  Article  Google Scholar 

  • Malik KA, Saxena PK. 1992a. Somatic embryogenesis and shoot regeneration from intact seedlings of Phaseolus acutifolius A., P. aureus (L.) Wilczek, P. coccineus L., and P. wrightii L. Plant Cell Reports 11:163–168

    CAS  Article  Google Scholar 

  • Malik KA, Saxena PK. 1992b. Thidiazuron induces high-frequency shoot regeneration in intact seedlings of pea (Pisum sativum), chickpea (Cicer arietinum) and lentil (Lens culinaris). Aust J Plant Physiol 19:731–740

    CAS  Article  Google Scholar 

  • Malik KA, Saxena PK. 1992c. Regeneration in Phaseolus vulgaris L.: high-frequency induction of direct shoot formation in intact seedlings by N6-benzylaminopurine and thidiazuron. Planta 186:384–389

    CAS  Article  Google Scholar 

  • Martin RC, Mok MC, Shaw G., Mok DWS. 1989. An enzyme mediating the conversion of zeatin to dihydrozeatin in Phaseolus embryos. Plant Physiol 90:1630–1635

    PubMed  CAS  Article  Google Scholar 

  • McKersie BD, Stinson RH. 1980. Effect of dehydration on leakage and membrane structure in Lotus corniculatus L. seeds. Plant Physiol 66:316–320

    PubMed  CAS  Google Scholar 

  • Mijatović M, Milijić S, Spasić M, Petrović R, Mitrović S. 1986. Morphology, biology and productivity in new cultivars of bird’s-foot trefoil Zora and Bokor. Arhiv za Poljoprivredne Nauke 47:149–155 (in Serbian)

    Google Scholar 

  • Motyka V, Faiss M, Strnad M, Kaminek M, Schmülling T. 1996. Changes in cytokinin content and cytokinin oxidase activity in response to derepression of ipt gene transcription in transgenic tobacco calli and plants. Plant Physiol 112:1035–1043

    PubMed  CAS  Google Scholar 

  • Murashige T, Skoog F. 1962. A revised medium for rapid growth and bioassays with tobacco tissue cultures. Physiol Plant 15:473–479

    CAS  Article  Google Scholar 

  • Nikolić R, Mitić N, Nešković M. 1997. Evaluation of agronomic traits in tissue culture-derived progeny of bird’s-foot trefoil. Plant Cell Tiss Org Cult 48:67–69

    Article  Google Scholar 

  • Nikolić R, Mitić N, Ninković S, Miljuš-Djukić J, Nešković M. 2003/4. Efficient genetic transformation of Lotus corniculatus L. and growth of transformed plants in field. Biol Plant 47:137–140

    Article  Google Scholar 

  • Rybczyński JJ, Badzian T. 1987. High regeneration potential of root segments of Lotus corniculatus L. seedlings on hormone free media. Plant Sci 51:239–244

    Article  Google Scholar 

  • Spíchal L, Rakova NYu, Riefler M, Mizuno T, Romanov GA, et al. 2004. Two cytokinin receptors of Arabidopsis thaliana, CRE1/AHK4 and AHK3, differ in their ligand specificity in a bacterial assay. Plant Cell Physiol 45:1299–1305

    PubMed  Article  Google Scholar 

  • Stirk WA, Gold JD, Novák O, Strnad M, van Staden J. 2005. Changes in endogenous cytokinins during germination and seedling establishment of Tagetes minuta L. 2005. Plant Growth Regul 47:1–7

    CAS  Article  Google Scholar 

  • Strnad M. 1997. The aromatic cytokinins. Physiol Plant 101:674–688

    CAS  Article  Google Scholar 

  • Victor JMR, Murch SJ, KrishnaRaj S, Saxena PK. 1999. Somatic embryogenesis and organogenesis in peanut: the role of thidiazuron and N6-benzylaminopurine in the induction of plant morphogenesis. Plant Growth Regul 28:9–15

    CAS  Article  Google Scholar 

  • Webb KJ, Watson EJ. 1991. Lotus corniculatus L. Morphological and cytological analysis of regenerants from three sources of tissue and selected progeny. Plant Cell Tiss Org Cult 25:27–33

    Article  Google Scholar 

  • Werner T, Motyka V, Strnad M, Schmülling T. 2001. Regulation of plant growth by cytokinin. Proc Natl Acad Sci USA 98:10487–10492

    PubMed  CAS  Article  Google Scholar 

Download references

Acknowledgments

The present work was supported by the Ministry of Science and Environment Protection of the Republic of Serbia (grant 3026).

Author information

Author notes

  1. Radomirka Nikolić & Nevena Mitić

    Present address: Institute for Biological Research “S. Stanković”, Belgrade University, Bul. Despota Stefana 142, 11060, Belgrade, Serbia

Authors and Affiliations

  1. Agricultural Research Institute “Serbia”, Center for Agricultural and Technological Research, 19000, Zaječar

    Radomirka Nikolić, Nevena Mitić & Rade Miletić

  2. Institute of Botany, Faculty of Biology and Institute for Biological Research “S. Stanković”, Belgrade University, Bul. Despota Stefana 142, 11060, Belgrade, Serbia

    Mirjana Nešković

Authors
  1. Radomirka Nikolić
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Nevena Mitić
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Rade Miletić
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Mirjana Nešković
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Radomirka Nikolić.

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Nikolić, R., Mitić, N., Miletić, R. et al. Effects of Cytokinins on In Vitro Seed Germination and Early Seedling Morphogenesis in Lotus corniculatus L.. J Plant Growth Regul 25, 187 (2006). https://doi.org/10.1007/s00344-005-0129-4

Download citation

  • Received:

  • Accepted:

  • Published:

  • DOI: https://doi.org/10.1007/s00344-005-0129-4

Keywords

  • Cytokinins
  • In vitro cultures
  • Seed germination
  • Shoot elongation
  • Root elongation
  • Shoot regeneration