Plant Cell, Tissue and Organ Culture (PCTOC)

, Volume 122, Issue 3, pp 751–757 | Cite as

Ethylene inhibitors improve efficiency of microspore embryogenesis in hexaploid triticale

  • Tobias Würschum
  • Matthew R. Tucker
  • Hans Peter Maurer
  • Willmar L. Leiser
Original Paper


Doubled haploid technology is an important tool in plant breeding and research, but routine application requires the establishment of efficient protocols. Microspore culture is an attractive approach although its efficiency is strongly dependent on the genotype. In this study we evaluated the effects of the three ethylene inhibitors aminooxyacetic acid, 2,5-norbornadiene, and silver thiosulphate (STS) on embryogenesis, regeneration rate and green plant rate in triticale microspore culture. Our results show that STS at 20 µM in particular had a positive effect on the embryogenesis rate, suggesting that for many genotypes ethylene accumulates in the culture vessel above a critical threshold where it becomes inhibitory for embryogenesis. In addition, STS also appears to positively influence the green plant rate. Taken together, our results show that the addition of ethylene inhibitors at defined concentrations can counteract the negative effect of ethylene, increase the embryogenesis rate and potentially also the green plant rate in microspore culture.


Triticale Microspore culture Doubled haploids Embryogenesis Ethylene inhibitors 



We greatly acknowledge the lab work of Alexandra Appel and Sylvia Kaiser.

Compliance with Ethical Standard

Ethical standard

The authors declare that the experiments comply with the current laws of Germany.

Conflict of interest

The authors declare that they have no conflict of interest.


  1. Asif M, Eudes F, Goyal A, Amundsen E, Randhawa H, Spaner D (2013) Organelle antioxidants improve microspore embryogenesis in wheat and triticale. In Vitro Cell Dev Biol Plant 49:489–497. doi: 10.1007/s11627-013-9514-z CrossRefGoogle Scholar
  2. Asif M, Eudes F, Randhawa H, Amundsen E, Spaner D (2014) Phytosulfokine alpha enhances microspore embryogenesis in both triticale and wheat. Plant Cell, Tissue Organ Cult 116:125–130. doi: 10.1007/s11240-013-0379-y CrossRefGoogle Scholar
  3. Beyer EM (1976) A potent inhibitor of ethylene action in plants. Plant Physiol 58:268–271PubMedCentralPubMedCrossRefGoogle Scholar
  4. Biddington NL (1992) The influence of ethylene in plant tissue culture. Plant Growth Regul 11:173–187CrossRefGoogle Scholar
  5. Biddington NL, Robinson TH (1991) Ethylene production during anther culture of Brussel sprouts (Brassica oleracea var. gemmifera) and its relationship with factors that affect embryo production. Plant Cell Tissue Cult 25:169–177Google Scholar
  6. Biddington NL, Sutherland RA, Robinson HT (1988) Silver nitrate increases embryo production in anther culture of Brussels sprouts. Ann Bot 62:181–185Google Scholar
  7. Castillo AM, Cistué L, Vallés MP, Soriano M (2009) Chromosome doubling in monocots. In: Touraev A, Forster BP, Mohan Jain S (eds) Advances in haploid production in higher plants. Springer, Berlin, pp 329–338CrossRefGoogle Scholar
  8. Cho UH, Kasha KJ (1989) Ethylene production and embryogenesis from anther cultures of barley (Hordeum vulgare). Plant Cell Rep 8:415–417PubMedCrossRefGoogle Scholar
  9. Dunwell JM (1979) Anther culture in Nicotiana tabacum: the role of culture vessel atmosphere in pollen embryo induction and growth. J Exp Bot 30:419–428CrossRefGoogle Scholar
  10. Eudes F, Amundsen E (2005) Isolated microspore culture of Canadian 6× triticale cultivars. Plant Cell, Tissue Organ Cult 82:233–241. doi: 10.1007/s11240-005-0867-9 CrossRefGoogle Scholar
  11. Ferrie AMR, Caswell KL (2011) Isolated microspore culture techniques and recent progress for haploid and doubled haploid plant production. Plant Cell, Tissue Organ Cult 104:301–309CrossRefGoogle Scholar
  12. Forster BP, Heberle-Bors E, Kasha KJ, Touraev A (2007) The resurgence of haploids in higher plants. Trends Plant Sci 12:368–375. doi: 10.1016/j.tplants.2007.06.007 PubMedCrossRefGoogle Scholar
  13. Hansen NJP, Andersen SB (1998) In vitro chromosome doubling with colchicine during microspore culture in wheat (Triticum aestivum L.). Euphytica 102:101–108CrossRefGoogle Scholar
  14. Hill SE, Stead AD, Nichols R (1987) Pollination-induced ethylene and production of 1-aminocyclopropane-1-carboxylic acid by pollen of Nicotiana tabacum cv. White Burley. J Plant Growth Regul 6:1–13CrossRefGoogle Scholar
  15. Horner M, McComb JA, McComb AJ, Street HE (1977) Ethylene production and plantlet formation by Nicotiana anthers cultured in the presence and abscence of charcoal. J Exp Bot 28:1365–1372Google Scholar
  16. Locke JM, Bryce JH, Morris PC (2000) Contrasting effects of ethylene perception and biosynthesis inhibitors on germination and seedling growth of barley (Hordeum vulgare L.). J Exp Bot 51:1843–1849PubMedCrossRefGoogle Scholar
  17. R Development Core Team (2010) R: a language and environment for statistical computing, R foundation for statistical computing.
  18. Sinha RK, Eudes F (2015) Dimethyl tyrosine conjugated peptide prevents oxidative damage and death of triticale and wheat microspores. Plant Cell, Tissue Organ Cult 122:227–237. doi: 10.1007/s11240-015-0763-x CrossRefGoogle Scholar
  19. Sisler EC, Pian A (1973) Effect of ethylene and cyclic olefins on tobacco leaves. Tob Sci 17:68–72Google Scholar
  20. Tiainen T (1992) The role of ethylene and reducing agents on anther culture response of tetraploid potato (Solanum tuberosum L.). Plant Cell Rep 10:604–607PubMedCrossRefGoogle Scholar
  21. Tiainen T (1996) Influence of ethylene in microspore embryogenesis. In: Jain SM, Sopory SK, Veilleux RE (eds) In vitro haploid production in higher plants, vol 1, pp 177–187Google Scholar
  22. Wedzony M, Forster BP, Zur I, Golemice E, Szechynska-Hebda M et al (2009) Progress in doubled haploid technology in higher plants. In: Touraev A, Forster BP, Mohan Jain S (eds) Advances in haploid production in higher plants. Springer, Berlin, pp 1–33CrossRefGoogle Scholar
  23. Williams J, Pink DAC, Biddington NL (1990) Effect of silver nitrate on long-term culture and regeneration of callus from Brassica oleracea var. gemmifera. Plant Cell, Tissue Organ Cult 21:61–66CrossRefGoogle Scholar
  24. Würschum T, Tucker MR, Reif JC, Maurer HP (2012) Improved efficiency of doubled haploid generation in hexaploid triticale by in vitro chromosome doubling. BMC Plant Biol 12:109. doi: 10.1186/1471-2229-12-109 PubMedCentralPubMedCrossRefGoogle Scholar
  25. Würschum T, Tucker MR, Maurer HP (2014) Stress treatments influence efficiency of microspore embryogenesis and green plant regeneration in hexaploid triticale (×Triticosecale Wittmack L.). In Vitro Cell Dev Biol Plant 50:143–148. doi: 10.1007/s11627-013-9539-3 CrossRefGoogle Scholar
  26. Yang SF, Hoffman NE (1984) Ethylene biosynthesis and its regulation in higher plants. Ann Rev Plant Physiol 35:155–189CrossRefGoogle Scholar
  27. Zheng MY, Liu W, Weng Y, Polle E, Konzak CF (2001) Culture of freshly isolated wheat (Triticum aestivum L.) microspores treated with inducer chemicals. Plant Cell Rep 20:685–690. doi: 10.1007/s00299-001-0393-0 CrossRefGoogle Scholar
  28. Żur I, Dubas E, Krzewska M, Janowiak F, Hura K et al (2014) Antioxidant activity and ROS tolerance in triticale (×Triticosecale Wittm.) anthers affect the efficiency of microspore embryogenesis. Plant Cell, Tissue Organ Cult 119:79–94. doi: 10.1007/s11240-014-0515-3 CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media Dordrecht 2015

Authors and Affiliations

  • Tobias Würschum
    • 1
  • Matthew R. Tucker
    • 2
  • Hans Peter Maurer
    • 1
  • Willmar L. Leiser
    • 1
  1. 1.State Plant Breeding InstituteUniversity of HohenheimStuttgartGermany
  2. 2.School of Agriculture, Food and Wine, Waite Research InstituteThe University of AdelaideGlen OsmondAustralia

Personalised recommendations