Skip to main content
SpringerLink
Log in

Growth Conditions in In Vitro Culture Can Induce Oxidative Stress in Mammillaria gracilis Tissues

  • Published:
Journal of Plant Growth Regulation Aims and scope Submit manuscript

Abstract

In vitro propagated plants of Mammillaria gracilis Pfeiff. (Cactaceae) develop calli without any exogenous growth regulators. This habituated tissue spontaneously regenerates morphologically normal as well as hyperhydric shoots. In this study, a possible involvement of activated oxygen metabolism in habituation and hyperhydricity in in vitro propagated plants of Mammillaria gracilis Pfeiff. (Cactaceae) was investigated. Significantly higher malondialdehyde (MDA) and carbonyl contents as well as hydrogen peroxide (H2O2) production were observed in habituated callus (HC), hyperhydric regenerated shoots (HS), and tumors (TT) in comparison to normal regenerated shoots (NS). Lipoxygenase (LOX) activity showed a similar trend, with a clear increase in activity in HC and HS. The activities of antioxidative enzymes, namely, peroxidase (POX), ascorbate peroxidase (APX), and catalase (CAT), were also higher in HC, HS, and TT, whereas an increase in superoxide dismutase (SOD) activity was observed in HC and HS. The majority of antioxidative isoenzymes were common to all cactus tissues, although a few tissue-specific bands were noticed. Significant decreases in phenylalanine ammonia lyase (PAL) activity, total phenolic content, and lignification were found in HS, HC, and TT in comparison to NS. Our results showed the appearance of a prominent oxidative stress in HC, HS, and TT as well as a strong induction of the antioxidant system indicating that activated oxygen metabolism could be involved in habituation and hyperhydricity and linked to the loss of tissue organization in M. gracilis.

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

Similar content being viewed by others

References

  • Aebi M (1984) Catalase in vitro. Meth Enzymol 105:121–126

    Article  PubMed  CAS  Google Scholar 

  • Axerold B, Cheesbrough TM, Laakso S (1981) Lipoxygenase from soybean. In: Lowenstein JM (ed) Methods in enzymology. Academic Press, New York, pp 441–451

    Google Scholar 

  • Banowetz GM, Dierksen KP, Azevedo MD, Stout R (2004) Microplate quantification of plant leaf superoxide dismutases. Anal Biochem 332:314–320

    Article  PubMed  CAS  Google Scholar 

  • Beauchamp C, Fridovich I (1971) Superoxide dismutase: improved assays and an assay applicable to acrylamide gels. Anal Biochem 44:276–287

    Article  PubMed  CAS  Google Scholar 

  • Bradford MM (1976) A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem 72:248–254

    Article  PubMed  CAS  Google Scholar 

  • Cassells AC, Curry RF (2001) Oxidative stress and physiological, epigenetic and genetic variability in plant tissue culture: implications for micropropagators and genetic engineers. Plant Cell Tiss Org Cult 64:145–157

    Article  CAS  Google Scholar 

  • Causevic A, Gentil MV, Delaunay A, El-Soud WA, Garcia Z, Pannetier C, Brignolas F, Hagege D, Maury S (2006) Relationship between DNA methylation and histone acetylation levels, cell redox and cell differentiation states in sugarbeet lines. Planta 224:812–827

    Article  PubMed  CAS  Google Scholar 

  • Chance B, Maehly AC (1955) Assay of catalases and peroxidases. In: Colowick SP, Kaplan NO (eds) Methods in enzymology. Academic Press, New York, pp 764–775

    Chapter  Google Scholar 

  • Chen J, Ziv M (2001) The effect of ancymidol on hyperhydricity, regeneration, starch and antioxidant enzymatic activities in liquid-cultured Narcissus. Plant Cell Rep 20:22–27

    Article  Google Scholar 

  • Dalle-Donne I, Rossi R, Giustarini D, Milzani A, Colombo R (2003) Protein carbonyl groups as biomarkers of oxidative stress. Clin Chim Acta 329:23–38

    Article  PubMed  CAS  Google Scholar 

  • Dewir YH, Chakrabarty D, Ali MB, Hahn EJ, Paek KY (2006) Lipid peroxidation and antioxidant enzyme activities of Euphorbia milli hyperhydric shoots. Environ Exp Bot 58:93–99

    Article  CAS  Google Scholar 

  • Fernandez-García N, Piqueras A, Olmos E (2008) Sub-cellular location of H2O2, peroxidases and pectin epitopes in control and hyperhydric shoots of carnation. Environ Exp Bot 62:168–175

    Article  Google Scholar 

  • Franck T, Kevers C, Penel C, Greppin H, Hausman JF, Gaspar T (1998) Reducing properties, and markers of lipid peroxidation in normal and hyperhydrating shoots of Prunus avium L. J Plant Physiol 153:339–346

    CAS  Google Scholar 

  • Franck T, Kevers C, Gaspar T, Dommes J, Deby C, Greimers R, Serteyn D, Deby-Dupont G (2004) Hyperhydricity of Prunus avium shoots cultured on gelrite: a controlled stress response. Plant Physiol Biochem 42:519–527

    Article  PubMed  CAS  Google Scholar 

  • Gaspar T, Kevers C, Bisbis B, Franck T, Crevecoeur M, Greppin H, Dommes J (2000) Loss of plant organogenic totipotency in the course of in vitro neoplastic progression. In Vitro Cell Dev Biol Plant 36:171–181

    Article  CAS  Google Scholar 

  • George EF (1996) Plant propagation by tissue culture. Part 2. The practice, 2nd edn. Exegetics Ltd., Edington, UK, p 1361

    Google Scholar 

  • Heath RL, Packer L (1968) Photoperoxidation in isolated chloroplasts. I–Kinetics and stoichiometry of fatty acid peroxidation. Arch Biochem Biophys 125:189–198

    Article  PubMed  CAS  Google Scholar 

  • Kevers C, Franck T, Strasser R, Dommes J, Gaspar T (2004) Hyperhydricity of micropropagated shoots: a typically stress-induced change of physiological state. Plant Cell Tiss Org Cult 77:181–191

    Article  Google Scholar 

  • Krsnik-Rasol M, Balen B (2001) Electrophoretic protein patterns and peroxidase activity related to morphogenesis in Mammillaria gracilis tissue culture. Acta Bot Croat 2:219–226

    Google Scholar 

  • Krsnik-Rasol M, Jelaska S (1991) Peroxidases in relation to differentiation and tumor transformation in plants. In: Lobrazewski J, Greppin H, Penel C, Gaspar T (eds) Biochemical, molecular and physiological aspects of plant peroxidases. University M. Curie-Sklodowska and University of Geneva, Lublin and Geneva, pp 373–382

    Google Scholar 

  • Krsnik-Rasol M, Muraja-Fras J (1993) Peroxidases as a morphogenesis marker in horseradish crown gall tumour. In: Welinder KG, Rasmunssen SK, Penel C, Greppin H (eds) Plant peroxidases: biochemistry and physiology. University of Geneva, Geneva, pp 423–428

    Google Scholar 

  • Laemmli UK (1970) Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227:680–685

    Article  PubMed  CAS  Google Scholar 

  • Levine RL, Williams JA, Stadtman ER, Shacter E (1994) Carbonyl assay for determination of oxidatively modified proteins. Method Enzymol 233:346–357

    Article  CAS  Google Scholar 

  • Malda G, Backhaus RA, Martin C (1999) Alterations in growth and crassulacean acid metabolism (CAM) activity of in vitro cultured cactus. Plant Cell Tiss Org Cult 58:1–9

    Article  CAS  Google Scholar 

  • Mittler R, Zilinskas BA (1993) Detection of ascorbate peroxidase activity in native gels by inhibition of the ascorbate-dependent reduction of nitroblue tetrazolium. Anal Biochem 212:540–546

    Article  PubMed  CAS  Google Scholar 

  • Mukherjee SP, Choudhari MA (1983) Implications of water stress-induced changes in the level of endogenous acsorbic acid and hydrogen peroxide in Vigna seedlings. Physiol Plant 58:166–170

    Article  CAS  Google Scholar 

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

    Article  CAS  Google Scholar 

  • Nakano Y, Asada K (1981) Hydrogen peroxide is scavenged by ascorbate peroxidase in spinach chloroplasts. Plant Cell Physiol 22:867–880

    CAS  Google Scholar 

  • Olmos E, Piqueras A, Martinez-Solano JR, Hellin E (1997) The subcellular localization of peroxidase and the implication of oxidative stress in hyperhydrated leaves of regenerated carnation shoots. Plant Sci 130:97–105

    Article  CAS  Google Scholar 

  • Perry PL, Ueno K, Shetty K (1999) Reversion to hyperhydration by addition of antibiotics to remove Pseudomonas in unhyperhydrated oregano tissue culture. Process Biochem 34:717–723

    Article  CAS  Google Scholar 

  • Poljuha D, Balen B, Bauer A, Ljubešić N, Krsnik-Rasol M (2003) Morphology and ultrastructure of Mammillaria gracilis (Cactaceae) in in vitro culture. Plant Cell Tiss Org Cult 75:117–123

    Article  Google Scholar 

  • Ros-Barceló A, Gómez-Ros LV, Ferrer MA, Hernandez JA (2006) The apoplastic antioxidant enzymatic system in the woodforming tissues of trees. Trees Struct Funct 20:145–156

    Google Scholar 

  • Saher S, Piqueras A, Hellin E, Olmos E (2004) Hyperhydricity in micropropagated carnation shoots: the role of oxidative stress. Physiol Plant 120:152–161

    Article  PubMed  CAS  Google Scholar 

  • Saher S, Fernández-García N, Piqueras A, Hellin E, Olmos E (2005) Reducing properties, energy efficiency and carbohydrate metabolism in hyperhydric and normal carnation shoots cultured in vitro: a hypoxia stress? Plant Physiol Biochem 43:573–582

    Article  PubMed  CAS  Google Scholar 

  • Sancho MA, Milrad de Fochetti S, Pliego F, Valpuesta V, Quesada MA (1996) Peroxidase activity and isoenzymes in the culture medium of NaCl adapted tomato suspenison cells. Plant Cell Tiss Org Cult 44:161–167

    Article  CAS  Google Scholar 

  • Saunders JA, McClure JW (1975) Phytochrome controlled phenylalanine ammonia lyase activity in Hordeum vulgare plastids. Phytochemistry 14:1285–1289

    Article  CAS  Google Scholar 

  • Slinkard K, Singleton VL (1997) Total phenol analyses: automation and comparison with manual methods. Am J Enol Viticult 28:49–55

    Google Scholar 

  • Surowka E, Karolewski P, Niewiadomska E, Libik M, Miszalski Z (2007) Antioxidative response of Mesembryanthemum crystallinum plants to exogenous SO2 application. Plant Sci 172:76–84

    Article  CAS  Google Scholar 

  • Woodbury WA, Spencer K, Stahlmann MA (1971) An improved procedure using ferricyanide for detecting catalase isozymes. Anal Biochem 44:301–305

    Article  PubMed  CAS  Google Scholar 

  • Ye SF, Zhou HY, Sun Y, Zou LY, Yu JQ (2006) Cinnamic acid causes oxidative stress in cucumber roots and promotes incidence of Fusarium wilt. Environ Exp Bot 56:255–262

    Article  CAS  Google Scholar 

Download references

Acknowledgments

The financial support of this work was provided by The Ministry of Science Education and Sports of the Republic of Croatia within projects 119-1191196-1200 (MKR) and 119-1191196-1202 (BPK).

Author information

Authors and Affiliations

  1. Department of Molecular Biology, Faculty of Science, University of Zagreb, Horvatovac 102a, HR-10000, Zagreb, Croatia

    Biljana Balen, Dubravko Pavoković & Marijana Krsnik-Rasol

  2. Department of Botany, Faculty of Science, University of Zagreb, Rooseveltov trg 6, HR-10000, Zagreb, Croatia

    Mirta Tkalec & Branka Pevalek-Kozlina

Authors
  1. Biljana Balen
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Mirta Tkalec
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Dubravko Pavoković
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Branka Pevalek-Kozlina
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Marijana Krsnik-Rasol
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Biljana Balen.

Additional information

B. Balen and M. Tkalec contributed equally to this work.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Balen, B., Tkalec, M., Pavoković, D. et al. Growth Conditions in In Vitro Culture Can Induce Oxidative Stress in Mammillaria gracilis Tissues. J Plant Growth Regul 28, 36–45 (2009). https://doi.org/10.1007/s00344-008-9072-5

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00344-008-9072-5

Keywords

Search

Navigation