Skip to main content
Log in

Efficient embryogenesis and regeneration in freshly isolated and cultured wheat (Triticum aestivum L.) microspores without stress pretreatment

  • Cell Biology and Morphogenesis
  • Published:
Plant Cell Reports Aims and scope Submit manuscript

Abstract

The major advantage of doubled haploids in plant breeding is the immediate achievement of complete homozygosity. Desired genotypes are thus fixed in one generation, reducing time and cost for cultivar or inbred development. Among the different technologies to produce doubled haploids, microspore embryogenesis is by far the most common. It usually requires reprogramming of microspores by stress such as cold, heat, and starvation, followed by embryo development under stress-free conditions. We report here the development of a simple and efficient isolated microspore culture system for producing doubled haploid wheat plants in a wide spectrum of genotypes, in which embryogenic microspores and embryos are formed without any apparent stress treatment. Microspores were isolated from fresh spikes in a nutrient-free medium by stirring and cultured in medium A2 in the dark at 25°C. Once embryogenic microspores were formed, ovaries and phytohormones were added directly to the cultures without changing the medium. The cultures were incubated in the dark at 25–27°C until the formation of embryos and then the embryos were transferred to regeneration medium. The regeneration frequency and percentage of green plants increased significantly using this protocol compared to the shed microspore culture method.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Subscribe and save

Springer+ Basic
EUR 32.99 /Month
  • Get 10 units per month
  • Download Article/Chapter or Ebook
  • 1 Unit = 1 Article or 1 Chapter
  • Cancel anytime
Subscribe now

Buy Now

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

Instant access to the full article PDF.

Similar content being viewed by others

References

  • Choo TM (1981a) Doubled haploids for estimating mean and variance of recombination values. Genetics 97:165–172

    PubMed  CAS  Google Scholar 

  • Choo TM (1981b) Doubled haploids for studying the inheritance of quantitative characters. Genetics 99:525–540

    PubMed  CAS  Google Scholar 

  • Dunwell JM (1985) Embryogenesis from pollen in vitro. In: Zaitlin P, Day P, Hollaender A (eds) Biotechnology in plant science. Academic Press, Orlando, pp 49–76

    Google Scholar 

  • Fischer C, Neuhaus G (1996) Influence of auxin on the establishment of bilateral symmetry in monocots. Plant J 9:659–669

    Article  CAS  Google Scholar 

  • Gustafson V, Baenziger PS, Wright MS, Stroup WW, Yen Y (1995) Isolated wheat microspore culture. Plant Cell Tiss Org Cult 42:207–213

    Article  Google Scholar 

  • Henry Y, de Buyser J (1989) Float culture of wheat anthers. Theor Appl Genet 60:77–79

    Google Scholar 

  • Herrmann RG, Feierabend J (1980) The presence of DNA in ribosome-deficient plastids of heat-bleached rye leaves. Eur J Biochem 104:603–609

    Article  PubMed  CAS  Google Scholar 

  • Hess WR, Blank-Huber M, Fieder B, Börner T, Rüdiger W (1992) Chlorophyl synthethase and chloroplast tRNAglu are present in heat-bleached, ribosome-deficient plastids. J Plant Physiol 139:427–430

    CAS  Google Scholar 

  • Hu TC, Ziauddin A, Simion E, Kasha KJ (1995) Isolated microspore culture of wheat (Triticum aestivum L.) in a defined media: I. Effects of pretreatment, isolation methods, and hormones. In Vitro Cell Dev Biol 31:79–83

    CAS  Google Scholar 

  • Hu TC, Kasha KJ (1997) Improvement of isolated microspore culture of wheat (Triticum aestivum L.) through ovary co-culture. Plant Cell Rep 16:520–525

    Article  CAS  Google Scholar 

  • Indrianto A, Heberle-Bors E, Touraev A (1999) Assessment of various stresses and carbohydrates for their effects on the induction of embryogenesis in isolated wheat microspores. Plant Sci 143:71–79

    Article  CAS  Google Scholar 

  • Indrianto A, Barinova J, Touraev A, Heberle-Bors E (2001) Tracking individual wheat microspores in vitro: identification of embryogenic microspores and body axis formation in the embryo. Planta 212:163–174

    Article  PubMed  CAS  Google Scholar 

  • Kasha KJ, Ziauddin A, Cho UH (1990) Haploids in Cereal improvement: anther and microspore culture. In: Gustafson JP (ed) Gene manipulation in plant improvement. Plenum Press, New York, pp 213–235

    Google Scholar 

  • Kumlehn J, Lörz H (1999) Monitoring sporophytic development of individual microspores of barley (Hordeum vulgare L.). In: Clement C, Audran CJ (eds) Anther and pollen, from biology to biotechnology. Springer Verlag, Berlin, pp 183–189

    Google Scholar 

  • Kyo M, Harada H (1986) Control of the developmental pathway of tobacco pollen in vitro. Planta 168:427–432

    Article  CAS  Google Scholar 

  • Lazar MD, Schaeffer GW, Baenziger PS (1985) The physical environment in relation to high frequency callus and plantlet development in anther culture of wheat (Triticum aestivum L.) cv. Chris. J Plant Physiol 121:103–109

    Google Scholar 

  • Letarte J, Simion E, Miner M, Kasha KJ (2005) Arabinogalactans and arabinogalactan-proteins induce embryogenesis in wheat (Triticum aestivum L.) microspore culture. Plant Cell Rep online first (doi: 10.1007/s00299-005-0013-5)

  • Liu W, Zheng MY, Polle E, Konzak CF (2002) Highly efficient doubled-haploid production in wheat (Triticum aestivum L.) via induced microspore embryogenesis. Crop Sci 42:686–692

    Article  Google Scholar 

  • Maluszynski M, Kasha KJ, Szarejko I (2003) Published doubled haploid protocols in plant species. In: Maluszynski M, Kasha KJ, Forster BP, Szarejko I (eds) Doubled haploid production in crop plants, a manual. Kluwer Academic Publishers, Dordrecht, The Netherlands, pp 309–335

    Google Scholar 

  • Mejza SJ, Morgant V, DiBona DE, Wong JR (1993) Plant regeneration from isolated microspores of Triticum aestivum. Plant Cell Rep 12:149–153

    Article  Google Scholar 

  • Patel M, Darvey NL, Marshall DR, Berry JO (2004) Optimization of culture conditions for improved plant regeneration efficiency from wheat microspore culture. Euphytica 140:197–204

    Article  CAS  Google Scholar 

  • Shariatpanahi ME, Bal U, Heberle-Bors E, Touraev A (2006) Stresses applied for the re-programming of plant microspores towards in vitro embryogenesis. Physiol Plant (in press)

  • Snape JW, Simpson E, Parker BB (1986) Criteria for the selection and use of doubled haploid systems in cereal breeding programmes. In: Horn W, Jensen CJ, Odenbach W, Schieder O (eds) Genetic manipulation in plant breeding. Walter de Gruiter, Berlin, New York, pp 217–229

    Google Scholar 

  • Tanaka M (1973) The effect of centrifugal treatment on the emergence of plantlets from cultured anther of tobacco. Jpn J Breed 23:171–174

    Google Scholar 

  • Touraev A, Indrianto A, Wratschko I, Vicente O, Heberle-Bors E (1996) Efficient microspore embryogenesis in wheat (Triticum aestivum L.) induced by starvation at high temperatures. Sex Plant Reprod 9:209–215

    Article  Google Scholar 

  • Touraev A, Vicente O, Heberle-Bors E (1997) Initiation of microspore embryogenesis by stress. Trends Plant Sci 2:297-302

    Article  Google Scholar 

  • Touraev A, Heberle-Bors E (1999) Microspore embryogenesis and in vitro pollen maturation in tobacco. In: Hall RD (ed) Methods in molecular biology: Vol. 111. Plant cell culture protocols. Humana Press Inc. Totowa, NJ, pp 281–291

    Chapter  Google Scholar 

  • Touraev A, Pfosser M, Heberle-Bors E (2001) The microspore: a haploid multipurpose cell. Adv Bot Res 35:53–109

    Google Scholar 

  • Tuvesson IKD, Öhlund RCV (1993) Plant regeneration through culture of isolated microspores of Triticum aestivum L. Plant Cell Tiss Org Cult 34:163–167

    Article  Google Scholar 

  • Vergne P, Delvallee I, Dumas C (1987) Rapid assessment of microspore and pollen development stage in wheat and maize using DAPI and membrane permealization. Stain Technol 62:299–304

    PubMed  CAS  Google Scholar 

  • Yoshida R, Kanno A, Sato T, Kameya T (1996a) Cool temperature-induced chlorosis in rice plants: I. Relationship between the induction and a disturbance of etioplast development. Plant Physiol 110:997–1005

    Article  PubMed  CAS  Google Scholar 

  • Yoshida R, Kanno A, Kameya T (1996b) Cool temperature-induced chlorosis in rice plants: II. Effects of cool temperature on the expression of plastid-encoded genes during shoot growth in darkness. Plant Phyiol 112:585–590

    CAS  Google Scholar 

  • 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

    Article  CAS  Google Scholar 

  • Zheng MY (2003a) Microspore culture in wheat (Triticum aestivum)—doubled haploid production via induced embryogenesis. Plant Cell Tiss Org Cult 73:213–230

    Article  CAS  Google Scholar 

  • Zheng MY, Liu W, Weng Y, Polle E, Konzak CF (2003b) Production of doubled haploids in wheat (Triticum aestivum L.) through microspore embryogenesis triggered by inducer chemicals. In: Maluszynski M, Kasha KJ, Forster BP, Szarejko I (eds) Doubled haploid production in crop plants, a manual. Kluwer Academic Publishers, Dordrecht, The Netherlands, pp 83–94

    Google Scholar 

  • Ziauddin A, Marsolais A, Simion E, Kasha KJ (1992) Improved plant regeneration from wheat anther and barley microspore culture using phenylacetic acid (PAA). Plant Cell Rep 11:489–498

    Article  CAS  Google Scholar 

Download references

Acknowledgements

Mehran E. Shariatpanahi thanks the Iranian government for a Ph.D scholarship. We are grateful to Seed and Plant Improvement Institute of Iran, Probsdorfer Saatzucht (Austria) and KWS AG (Germany) for providing seeds and Fatima Touraeva for plant care.

Author information

Authors and Affiliations

Authors

Corresponding author

Correspondence to Alisher Touraev.

Additional information

Communicated by W. Harwood

Rights and permissions

Reprints and permissions

About this article

Cite this article

Shariatpanahi, M.E., Belogradova, K., Hessamvaziri, L. et al. Efficient embryogenesis and regeneration in freshly isolated and cultured wheat (Triticum aestivum L.) microspores without stress pretreatment. Plant Cell Rep 25, 1294–1299 (2006). https://doi.org/10.1007/s00299-006-0205-7

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00299-006-0205-7

Key words

Navigation