Plant Growth Regulation

, Volume 77, Issue 2, pp 125–132 | Cite as

Cytokinin profiling of long-term in vitro pea (Pisum sativum L.) shoot cultures

  • Lenka Plačková
  • Jakub Hrdlička
  • Iva Smýkalová
  • Magdalena Cvečková
  • Ondřej Novák
  • Miroslav Griga
  • Karel Doležal
Original paper

Abstract

Forty-seven endogenous cytokinin metabolites were determined in samples of long-term in vitro multiple shoot cultures of pea (Pisum sativum L.). Significant differences were found in the levels of all cytokinin groups except cis-zeatin and some of its derivatives. Old cultures (30 or 10 years old) contained larger total and active cytokinin pools than freshly prepared cultures. Long-term shoot cultures maintained on cytokinin-supplemented media initially exhibit gradually increasing concentrations of endogenous cytokinins that subsequently stabilise at a high level, which can make it more difficult to induce rooting later on. This is consistent with previous findings concerning the growth and development of plantlets derived from long-term cultures and the generally accepted role of cytokinins during root development. The results presented herein demonstrate the utility of long-term pea cultures as model systems for studying the influence of long-term cultivation on genetic stability, and suggest ways of circumventing the rooting problems associated with such cultures.

Keywords

Cytokinins Pea Quantification UHPLC–MS/MS Long-term culture in vitro 

Abbreviations

BAP

6-Benzylaminopurine

cZ

cis-Zeatin

DHZ

Dihydrozeatin

*9G

9-β-d-Glucopyranosyl derivative

*5′MP

5′-Monophosphate derivative

*OG

O-β-d-Glucopyranosyl derivative

tZ

trans-Zeatin

UHPLC–ESI(+)–MS/MS

Ultra high-performance liquid chromatography–electrospray tandem mass spectrometry

Supplementary material

10725_2015_44_MOESM1_ESM.docx (58 kb)
Supplementary material 1 (DOCX 57 kb)

References

  1. Aremu AO, Plačková L, Bairu MV, Novák O, Szüčová L, Doležal K, Finnie JF, Van Staden J (2012) Topolins: a panacea to plant tissue culture challenges? Plant Cell Tissue Organ Cult 108:1–16CrossRefGoogle Scholar
  2. Aremu AO, Bairu MV, Szüčová L, Doležal K, Finnie JF, Van Staden J (2013) Genetic fidelity in tissue-cultured ‘Williams’ bananas—the effect of high concentration of topolins and benzyladenine. Sci Hortic 161:324–327CrossRefGoogle Scholar
  3. Aremu AO, Bairu MV, Doležal K, Finnie JF, Van Staden J (2014) Endogenous cytokinin profiles of tissue-cultured and acclimatized ‘Williams’ bananas subjected to different aromatic cytokinin treatments. Plant Sci 214:88–98CrossRefPubMedGoogle Scholar
  4. Bairu MW, Novák O, Doležal K, Van Staden J (2011) Changes in endogenous cytokinin profiles in micropropagated Harpagophytum procumbens in relation to shoot-tip necrosis and cytokinin treatments. Plant Growth Regul 63:105–114CrossRefGoogle Scholar
  5. Bajguz A, Piotrowska A (2009) Conjugates of auxin and cytokinin. Phytochemistry 70:957–969CrossRefPubMedGoogle Scholar
  6. Bieleski RL (1964) The problem of halting enzyme action when extracting plant tissues. Anal Biochem 9:431–442CrossRefPubMedGoogle Scholar
  7. Del Bianco M, Giustini L, Sabatini S (2013) Spatiotemporal changes in the role of cytokinin during root development. New Phytol 199:324–338CrossRefPubMedGoogle Scholar
  8. Doležal K, Popa I, Kryštof V, Spíchal L, Fojtíková M, Holub J, Lenobel R, Schmülling T, Strnad M (2006) Preparation and biological activity of 6-benzylaminopurine derivatives in plants and human cancer cells. Bioorg Med Chem 14:875–884CrossRefPubMedGoogle Scholar
  9. Doležal K, Popa I, Hauserová E, Spíchal L, Chakrabarty K, Novák O, Kryštof V, Voller J, Holub J, Strnad M (2007) Preparation, biological activity and endogenous occurrence of N6-benzyladenosines. Bioorg Med Chem 15:3737–3747CrossRefPubMedGoogle Scholar
  10. Ezhova TA, Bagrova AM, Gostimski SA (1995) Cell selection as a possible reason for the specificity of somaclonal variation in pea. Plant Breed 114:520–524CrossRefGoogle Scholar
  11. Faiss M, Zalubilová J, Strnad M, Schmulling T (1997) Conditional transgenic expression of the ipt gene indicates a function for cytokinins in paracrine signaling in whole tobacco plants. Plant J 12:401–415CrossRefPubMedGoogle Scholar
  12. Fletcher PJ (1994) In vitro long-term storage of asparagus. N Z J Crop Hort Sci 22:351–359CrossRefGoogle Scholar
  13. Gajdošová S, Spíchal L, Kamínek M, Hoyerová K, Novák O, Dobrev PI, Galuszka P, Klíma P, Gaudinova A, Žižková E, Hanuš J, Dančák M, Trávníček B, Pešek B, Krupička M, Vaňková R, Strnad M, Motyka V (2011) Distribution, biological activities, metabolism, and the conceivable function of cis-zeatin-type cytokinins in plants. J Exp Bot 62:2827–2840CrossRefPubMedGoogle Scholar
  14. Gamborg OL, Miller RA, Ojima K (1968) Nutrient requirements of suspension cultures of soyabean root cells. Exp Cell Res 50:151–158CrossRefPubMedGoogle Scholar
  15. George PS, Ravishankar GA (1997) In vitro multiplication of Vanilla planifolia using axillary bud explants. Plant Cell Rep 16:490–494Google Scholar
  16. Gostimskii SA, Bagrova AM, Ezhova TA (1995) Discovery and cytogenetic analysis of the variability in plants regenerated from tissue-culture of pea (Pisum sativum L.). Dokl Akad Nauk SSSR 283:1007–1011Google Scholar
  17. Griga M, Novák FJ (1990) Pea (Pisum sativum L.). In: Bajaj YPS (ed) Biotechnology in agriculture and forestry, legumes and oil seed crops, vol 10. Springer, Berlin, pp 65–99Google Scholar
  18. Griga M, Stejskal J (1994) Micropropagation of pea (Pisum sativum L.)—in vitro system and its practical applications. In: Lumsden PJ, Nicholas JR, Davies WJ (eds) Physiology, growth and development of plants in culture. Kluwer Academic Publishers, Dordrecht, pp 278–283CrossRefGoogle Scholar
  19. Griga M, Tejklová E, Novák FJ (1984) Hormonal regulation of growth of pea (Pisum sativum L.) shoot apices in in vitro culture. Rost Výr 30:523–530 (in Czech)Google Scholar
  20. Griga M, Tejklová E, Novák FJ, Kubaláková M (1986) In vitro clonal propagation of Pisum sativum L. Plant Cell Tissue Organ Cult 6:95–104CrossRefGoogle Scholar
  21. Inpuay K, Arthipatjaporn A, Te-chato S (2012) Assessment of genetic instability of regenerated plantlets from long-term culture of oil palm through SSE formation by SSR marker. Int J Agric Technol 8:585–595Google Scholar
  22. Jevremović S, Trifunović M, Nikolić M, Subotić A, Radojević L (2006) Clonal fidelity of Chrysanthemum regenerated from long term cultures. Genetika 38:243–249CrossRefGoogle Scholar
  23. Jevremović S, Subotić D, Mijlković D, Trifunović M, Petrić M, Cingel A (2012) Clonal fidelity of Chrasanthemum cultivars after long term micropropagation by stem segment culture. Acta Hort (ISHS) 961:211–216CrossRefGoogle Scholar
  24. Kubaláková M, Tejklová E, Griga M (1988) Some factors affecting Root formation on in vitro regenerated pea shoots. Biol Plant 30:179–184CrossRefGoogle Scholar
  25. Kumar PS, Mathur VL (2004) Chromosomal instability in callus culture of Pisum sativum. Plant Cell Tissue Organ Cult 78:267–271CrossRefGoogle Scholar
  26. Lakshmanan V, Venkataramareddy SR, Neelwarne B (2007) Molecular analysis of genetic stability in long-term micropropagated shoots of banana using RAPD and ISSR markers. Electr J Biotechnol 10:106–113Google Scholar
  27. Lebeda A, Švábová L (2010) In vitro screening methods for assessing of plant disease resistance. Mass screening techniques for selecting crops resistant to diseases. IAEA, Vienna, pp 5–45Google Scholar
  28. Malá J, Máchová P, Cvrčková H, Karady M, Novák O, Mikulík J, Dostál J, Strnad M, Doležal K (2013) The role of cytokinins during micropropagation of wych elm. Biol Plant 57(1):174–178CrossRefGoogle Scholar
  29. Moubayidin L, Perilli S, Dello Ioio R, Di Mambro R, Costantino P, Sabatini S (2010) The rate of cell differentiation controls the Arabidopsis root meristem growth phase. Curr Biol 20:1138–1143CrossRefPubMedGoogle Scholar
  30. Murashige T, Skoog F (1962) A revised medium for rapid growth and bioassays with tobacco tissue cultures. Physiol Plant 15:472–497CrossRefGoogle Scholar
  31. Ngezahayo F, Liu B (2014) Axillary bud proliferation approach for plant biodiversity conservation and restoration. Intern J Biodivers. doi:10.1155/2014/727025 Google Scholar
  32. Nordström A, Tarkowski P, Tarkowská D, Norbaek R, Åstot C, Doležal K, Sandberg G (2004) Auxin regulation of cytokinin biosynthesis in Arabidopsis thaliana: a factor of potential importance for auxin–cytokinin-regulated development. PNAS 101:8039–8044PubMedCentralCrossRefPubMedGoogle Scholar
  33. Novák O, Hauserová E, Amakorová P, Doležal K, Strnad M (2008) Cytokinin profiling in plant tissues using ultra-performance liquid chromatography–electrospray tandem mass spectrometry. Phytochemistry 69:2214–2224CrossRefPubMedGoogle Scholar
  34. Rival A, Tregear J, Jaligot E, Morcillo F, Aberlenc F, Billotte N, Richaud F, Beulé T, Borgel A, Duval Y (2003) Biotechnology of the oil palm (Elaeis guineensis Jacq). In: Jaiwal Pawan K, Singh Rana P (eds) Plant genetic engineering. Improvement of commercial plants–II. Sci Tech Publishing, Houston, pp 261–318Google Scholar
  35. Skoog F, Armstrong DJ (1970) Cytokinins. Ann. Rev Plant Physiol 21:359–384CrossRefGoogle Scholar
  36. Skoog F, Miller CO (1957) Chemical regulation of growth and organ formation in plant tissues cultured in vitro. Symp Soc Exp Biol 11:118–131PubMedGoogle Scholar
  37. Smýkal P, Valledor L, Rodriguez R, Griga M (2007) Assessment of genetic and epigenetic stability in long-term in vitro shoot culture of pea (Pisum sativum L.). Plant Cell Rep 26:1985–1998CrossRefPubMedGoogle Scholar
  38. Sreedhar RV, Venkatachalam L, Bhagyalaksmi N (2007) Genetic fidelity of long-term micropropagated shoot cultures of vanilla (Vanilla planifolia Andrews) as assessed by molecular markers. Biotechnol J 2:1–7CrossRefGoogle Scholar
  39. Strnad M (1997) The aromatic cytokinins. Physiol Plantarum 101:674–688CrossRefGoogle Scholar
  40. Švábová L, Griga M (2008) The effect of cocultivation treatments on transformation efficiency in pea (Pisum sativum L.). Plant Cell Tissue Organ Cult 95:293–304CrossRefGoogle Scholar
  41. Švábová L, Lebeda A (2005) In vitro selection for improved plant resistance to toxin-producing pathogens. J Phytopathol 153:52–64CrossRefGoogle Scholar
  42. Švábová L, Smýkal P, Griga M, Ondřej V (2005) Agrobacterium-mediated transformation of Pisum sativum in vitro and in vivo. Biol Plant 49:361–370CrossRefGoogle Scholar
  43. Švábová L, Lebeda A, Kitner M, Sedlářová M, Petřivalský M, Dostálová R, Ondřej M, Horáček J, Smýkalová I, Griga M (2011) Comparison of the effects of Fusarium solani filtrates in vitro and in vivo on morphological characteristics and peroxidase activity in pea cultivars with different susceptibility. J Plant Pathol 93:19–30Google Scholar
  44. Szüčová L, Spíchal L, Doležal K, Zatloukal M, Greplová J, Galuszka P, Kryštof V, Voller J, Popa I, Massino FJ, Jørgensen J-E, Strnad M (2009) Synthesis, characterization and biological activity of ring-substituted 6-benzylamino-9-tetrahydropyran-2-yl and 9-tetrahydrofuran-2-ylpurine derivatives. Bioorg Med Chem 17:1938–1947CrossRefPubMedGoogle Scholar
  45. Werbrouck S, Van der Jeugt B, Dewitte W, Prinsen E, Van Onckelen H, Debergh P (1995) The metabolism of benzyladenine in Spathiphyllum floribundum ‘Schott Petite’ in relation to acclimatisation problems. Plant Cell Rep 14:662–665CrossRefPubMedGoogle Scholar
  46. Werbrouck SPO, Strnad M, Van Onckelen HA, Debergh PC (1996) Meta-topolin, an alternative to benzyladenine in tissue culture? Physiol Plant 98:291–297CrossRefGoogle Scholar
  47. Zatloukal M, Gemrotová M, Doležal K, Havlíček L, Spíchal L, Strnad M (2008) Novel potent inhibitors of A. thaliana cytokinin oxidase/dehydrogenase. Bioorg Med Chem. 16:9268–9275CrossRefPubMedGoogle Scholar

Copyright information

© Springer Science+Business Media Dordrecht 2015

Authors and Affiliations

  • Lenka Plačková
    • 1
  • Jakub Hrdlička
    • 1
  • Iva Smýkalová
    • 2
  • Magdalena Cvečková
    • 2
  • Ondřej Novák
    • 1
  • Miroslav Griga
    • 2
  • Karel Doležal
    • 1
  1. 1.Laboratory of Growth Regulators & Department of Chemical Biology and Genetics, Faculty of Science, Centre of the Region Haná for Biotechnological and Agricultural ResearchPalacký University and Institute of Experimental Botany ASCROlomoucCzech Republic
  2. 2.Plant Biotechnology DepartmentAGRITEC Plant Research Ltd.ŠumperkCzech Republic

Personalised recommendations