Sexual Plant Reproduction

, Volume 9, Issue 4, pp 209–215 | Cite as

Efficient microspore embryogenesis in wheat (Triticum aestivum L.) induced by starvation at high temperature

  • A. TouraevEmail author
  • A. Indrianto
  • I. Wratschko
  • O. Vicente
  • E. Heberle-Bors
Original Paper


We have established an efficient method to induce embryo formation from isolated wheat (Triticum aestivum L.) microspores. Culture of excised anthers under starvation and heat shock conditions induced the formation of embryogenic microspores at high frequency in nine Austrian winter wheat genotypes, including cultivars that had been considered as recalcitrant in anther culture. Percoll gradient centrifugation of the mechanically isolated microspores allowed us to obtain homogeneous populations of embryogenic microspores in all genotypes which, after transfer to a rich medium containing immature ovaries for conditioning, divided and produced globular embryos. Thousands of embryos were produced in one petri dish. Many of these embryos developed into plantlets after transfer to a solid medium without ovaries.

Key words

Triticum aestivum L. Microspore embryogenesis Starvation Heat shock Haploid production 


Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.


  1. Benito Moreno RM, Macke F, Hauser M-T, Alwen A, Heberle-Bors E (1988) Sporophytes and male gametophytes from in vitro cultured, immature tobacco pollen. In: Cresti M, Gori P, Pacini E (eds) Sexual reproduction in higher plants. Springer, Berlin Heidelberg New York, pp 137–142Google Scholar
  2. Chu CC, Hill RD, Brule-Babel AL (1990) High frequency of pollen embryoid formation and plant regeneration inTriticum aestivum L. on monosaccharide containing media. Plant Sci 66:255–262Google Scholar
  3. Datta SK, Wenzel G (1987) Isolated microspore derived plant formation via embryogenesis inTriticum aestivum. Plant Sci 48: 49–54Google Scholar
  4. Gamborg OL (1970) The effects of amino acids and ammonium on the growth of plant cells in suspension culture. Plant Physiol 45:372–375Google Scholar
  5. Garrido D, Charvat B, Benito Moreno RM, Alwen A, Vicente O, Heberle-Bors E (1991) Pollen culture for haploid plant production in tobacco. In: Negrutiu I, Gharti-Chhetri G (eds) A laboratory guide for cellular and molecular plant biology. Birkhäuser, Basel, pp 59–69Google Scholar
  6. Heberle-Bors E (1985) In vitro haploid formation from pollen: a critical review. Theor Appl Genet 71:361–374Google Scholar
  7. Heberle-Bors E (1989) Isolated pollen culture in tobacco: plant reproductive development in a nutshell. Sex Plant Reprod 2:1–10Google Scholar
  8. Hoekstra S, Zijderveld MH van, Louwerse JD, Heidekamp F, Mark F van der (1992) Anther and microspore culture ofHordeum vulgare L. cv. Igri. Plant Sci 86:89–96Google Scholar
  9. Hoekstra S, Zijderveld MN van, Heidekamp F, Mark F van der (1993) Microspore culture ofHordeum vulgare L.: the influence of density and osmolality. Plant Cell Rep 12:661–665Google Scholar
  10. Hu H, Huang B (1987) Application of pollen-derived plants to crop improvement. Int Rev Cytol 107:293–311Google Scholar
  11. Hu TC, Ziauddin A, Simion E, Kasha KJ (1995) Isolated microspore culture of wheat (Triticum aestivum L.) in a defined media. 1. Effects of pretreatments, isolation methods, and hormones. In Vitro Cell Dev Biol (Plant) 31:79–83Google Scholar
  12. Huang B (1987) Effects of incubation temperature on microspore callus production and plant regeneration in wheat anther cultures. Plant Cell Tissue Organ Cult 9:45–48Google Scholar
  13. Huang B (1992) Genetic manipulation of microspores and microspore derived embryos. In Vitro Cell Dev Biol 28:53–58Google Scholar
  14. Jähne A, Becker D, Brettschneider R, Lörz H (1994) Regeneration of transgenic, microspore-derived, fertile barley. Theor Appl Genet 89:525–533Google Scholar
  15. Köhler F, Wenzel G (1985) Regeneration of isolated barley microspores in conditioned media and trials to characterize the responsible factor. J Plant Physiol 121:181–191Google Scholar
  16. Kuhlmann U, Forough-Wehr B, Graner A, Wenzel G (1991) Improved culture system for microspores of barley to become a target for DNA uptake. Plant Breed 107:165–168Google Scholar
  17. Kyo M, Harada H (1986) Control of the developmental pathway of tobacco pollen in vitro. Planta 168:427–432Google Scholar
  18. Li HC, Qureshi J, Kariba KK (1988) The influence of different temperature treatments on anther culture response of spring wheat (Triticum aestivum L.). Plant Sci 57:55–61Google Scholar
  19. Lichter R (1982) Induction of haploid plants from isolated pollen ofBrassica napus L. Z Pflanzenphysiol 105:427–434Google Scholar
  20. Lichter R (1989) Efficient yield of embryoids by culture of isolated microspores of differentBrassicaceae species. Plant Breed 103:119–123Google Scholar
  21. Löschenberger F, Heberle-Bors E (1992) Anther culture responsiveness of Austrian winter wheat (Triticum aestivum L.) cultivars. Die Bodenkultur 43:115–122Google Scholar
  22. Marsolais AA, Seguin-Swartz G, Kasha KJ (1984) The influence of anther cold pretreatments and donor plant genotypes on in vitro androgenesis in wheat (Triticum aestivum). Plant Cell Tissue Organ Cult 3:69–79Google Scholar
  23. Mejza SJ, Morgant V, DiBona D, Wong JR (1993) Plant regeneration from isolated microspores ofTriticum aestivum. Plant Cell Rep 12:149–153Google Scholar
  24. Mordhorst AP, Lörz H (1993) Embryogenesis and development of isolated barley (Hordeum vulgare L.) microspores are influenced by the amount and composition of nitrogen sources in culture. J Plant Physiol 142:485–492Google Scholar
  25. Morrison RA, Evans D (1988) Haploid plants from tissue culture: new plant varieties in a shortened time frame. Biotechnology 6:684–690Google Scholar
  26. Ogawa T, Fukuoka H, Ohkawa Y (1994) Induction of cell division of isolated pollen grains by sugar starvation in rice. Breed Sci Jpn 44:75–77Google Scholar
  27. Olsen FL (1991) Isolation and cultivation of embryogenic microspores from barley (Hordeum vulgare L.). Hereditas 115:255–266Google Scholar
  28. Ouyang JW, Zhou SM, Jia SE (1983) The response of anther culture to culture temperature inTriticum aestivum. Theor Appl Genet 66:101–109Google Scholar
  29. Ouyang JW, He DG, Feng GH, Jia SE (1987) The response of anther culture to culture temperature varies with growth conditions of anther-donor plants. Plant Sci 49:145–148Google Scholar
  30. Pechan PM, Keller WA (1988) Identification of potentially embryogenic microspores inBrassica napus. Physiol Plant 74: 377–384Google Scholar
  31. Stoger E, Fink C, Pfosser M, Heberle-Bors E (1995) Plant transformation by particle bombardment of embryogenic pollen. Plant Cell Rep 14:273–278Google Scholar
  32. Sunderland N, Xu ZH (1982) Shed pollen culture inHordeum vulgare. J Exp Bot 33:1086–1095Google Scholar
  33. Takahata Y, Keller AW (1991) High frequency embryogenesis and plant regeneration in isolated microspore culture ofBrassica oleracea L. Plant Sci 74:235–242Google Scholar
  34. Telmer CA, Simmonds DH, Newcomb W (1992) Determination of developmental stage to obtain high frequencies of embryogenic microspores inBrassica napus. Physiol Plant 84: 417–424Google Scholar
  35. Touraev A, Ilham A, Vicente O, Heberle-Bors E (1996a) Stress-induced microspore embryogenesis in tobacco: an optimized system for molecular studies. Plant Cell Rep 15:561–565Google Scholar
  36. Touraev A, Pfosser M, Vicente O, Heberle-Bors E (1996b) Stress as the major signal controlling the developmental fate of tobacco microspores; towards a unified model of induction of microspore/pollen embryogenesis. Planta (in press)Google Scholar
  37. Tuevesson IKD, Ohlund RCV (1993) Plant regeneration through culture of isolated microspores ofTriticum aestivum L. Plant Cell Tissue Organ Cult 34:163–167Google Scholar
  38. Vergne P, Delvallee I, Dumas C (1987) Rapid assessement of microspore and pollen development stage in wheat and maize using DAPI and membrane permealization. Stain Technol 62:299–304Google Scholar
  39. Vicente O, Benito-Moreno RM, Heberle-Bors E (1991) Pollen cultures as a tool to study plant development. Cell Biol Rev 25:295–306Google Scholar
  40. Xu ZH, Huang B, Sunderland N (1981) Culture of barley anthers in conditioned media. J Exp Bot 32:767–778Google Scholar
  41. Ziauddin A, Simion E, Kasha KJ (1990) Improved plant regeneration from shed microspore culture in barley (Hordeum vulgare L.) cv. Igri Plant Cell Rep 9:69–72Google Scholar

Copyright information

© Springer-Verlag 1996

Authors and Affiliations

  • A. Touraev
    • 1
    Email author
  • A. Indrianto
    • 1
  • I. Wratschko
    • 1
  • O. Vicente
    • 1
  • E. Heberle-Bors
    • 1
  1. 1.Institute of Microbiology and GeneticsVienna BiocenterViennaAustria

Personalised recommendations