Efficiency of three haplomethods in durum wheat (Triticum turgidum subsp. durum Desf.): isolated microspore culture, gynogenesis and wheat × maize crosses

https://doi.org/10.17221/188/2017-CJGPBCitation:Slama-Ayed O., Bouhaouel I., Ayed S., De Buyser J., Picard E., Slim Amara H. (2019): Efficiency of three haplomethods in durum wheat (Triticum turgidum subsp. durum Desf.): isolated microspore culture, gynogenesis and wheat × maize crosses. Czech J. Genet. Plant Breed., 55: 101-109.
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This study presents the first report comparing the efficiency of microspore culture, gynogenesis and durum wheat × maize crosses for haploid plant production from three durum wheat genotypes (Razzek, Karim and Jneh Khotifa). The results showed that the best induction, calli or embryos formation and plant regeneration rates for the three genotypes were obtained with gynogenesis (47.2, 7.6, 0.8%), followed by interspecific crosses (33.1, 1.7, 0.4%) and isolated microspore culture (8.2, 0.05, 0.01%). Interestingly, all plants regenerated by gynogenesis and durum wheat × maize crosses were green whereas all plants obtained by isolated microspore culture were albino. In the haploid production system, all steps of the process are important for the three methods. The critical steps that have greatly reduced the number of regenerated haploid plants were induction, embryogenesis and regeneration for microspore culture, forming and regeneration of calli or embryo and haploid regeneration for interspecific crosses and gynogenesis. Genotypes with good capacity of induction have not necessarily a good capacity of haploid plantlets regeneration and vice-versa. However, calli or embryos formation seems to be an indicator of the haploid production. Overall, Razzek showed a good ability to produce haploids using the three methods. Each haplomethod showed a specic advantage. Although gynogenesis is the less used method for durum wheat, it has proved to be a successful approach for green haploid plant production.


Ayed S., Slama-Ayed O., Slim-Amara H. (2011a): Effect of 2,4-dichlorophenoxyacetic acid and nitrate silver on the efficiency of haploid production in durum wheat × maize crosses. International Journal of Plant Breeding, 5: 101–105.
Ayed S., Slama-Ayed O., Jaime A.T., Slim-Amara H. (2011b): Effect of different factors on haploid production through embryo rescue in durum wheat × maize crosses. International Journal of Plant Breeding, 5: 118–121.
Bal U., Shariatpanahi M.E., Castro A.J., Emery D., Clément C., Dehestani-Ardakani M., Mozaffari K., Touraev A. (2012): Pseudo-embryogenic structures in anther and isolated microspore cultures in vitro: a cautionary guide. Czech Journal of Genetics and Plant Breeding, 48, 51-60  https://doi.org/10.17221/176/2011-CJGPB
BARCLAY I. R. (1975): High frequencies of haploid production in wheat (Triticum aestivum) by chromosome elimination. Nature, 256, 410-411  https://doi.org/10.1038/256410a0
Campion B., Alloni C. (1990): Induction of haploid plants in onion (Allium cepa L.) by in vitro culture of unpollinated ovules. Plant Cell, Tissue and Organ Culture, 20, 1-6  https://doi.org/10.1007/BF00034750
Castillo A.M., Vallés M.P., Cistué L. (2000): Comparison of anther and isolated microspore cultures in barley. Effects of culture density and regeneration medium. Euphytica, 113: 1–8. https://doi.org/10.1023/A:1003937530907
Castillo Ana M., Sánchez-Díaz Rosa A., Vallés María P. (2015): Effect of ovary induction on bread wheat anther culture: ovary genotype and developmental stage, and candidate gene association. Frontiers in Plant Science, 6, -  https://doi.org/10.3389/fpls.2015.00402
Chaudhary H.K., Mahato A., Kaila V., Rather S.A., Tayeng T. (2015): Dihaploid induction in tetraploid durum wheat (Triticum durum L.) using pollen of Imperata cylindrica. Czech Journal of Genetics and Plant Breeding, 4: 142–147.
Cherkaoui S., Lamsaouri O., Chlyah A., Chlyah H. (2000): Durum wheat x maize crosses for haploid wheat production: Influence of parental genotypes and various experimental factors. Plant Breeding, 119, 31-36  https://doi.org/10.1046/j.1439-0523.2000.00433.x
Chu C.C., Hill R.D., Brule-Babel L. (1990): High frequency of pollen embryoid formation and plant regeneration in Triticum aestivum L. on monosaccharide containing media. Plant Science, 66, 255-262  https://doi.org/10.1016/0168-9452(90)90211-6
Cistué L., Soriano M., Castillo A. M., Vallés M. P., Sanz J. M., Echávarri B. (2006): Production of doubled haploids in durum wheat (Triticum turgidum L.) through isolated microspore culture. Plant Cell Reports, 25, 257-264  https://doi.org/10.1007/s00299-005-0047-8
Cistué Luis, Romagosa I., Batlle F., Echávarri B. (2009): Improvements in the production of doubled haploids in durum wheat (Triticum turgidum L.) through isolated microspore culture. Plant Cell Reports, 28, 727-735  https://doi.org/10.1007/s00299-009-0690-6
De Buyser J., Touraine P., Jaiti F., Picard E. (2002): Haplodiploidization by Isolated Microspore Culture from in vitro Wheat. Plant Biotechnology: Laboratory techniques. Montreal, Livre Robert Haïcour, AUF: 257−273. (in French)
Gamborg Oluf L., Eveleigh D. E. (1968): Culture methods and detection of glucanases in suspension cultures of wheat and barley. Canadian Journal of Biochemistry, 46, 417-421  https://doi.org/10.1139/o68-063
Garc�a-llamas C., Mart�n A., Ballesteros J. (2004): Differences among auxin treatments on haploid production in durum wheat � maize crosses. Plant Cell Reports, 23, -  https://doi.org/10.1007/s00299-004-0786-y
Germanà Maria Antonietta (2011): Gametic embryogenesis and haploid technology as valuable support to plant breeding. Plant Cell Reports, 30, 839-857  https://doi.org/10.1007/s00299-011-1061-7
Gürel S., Gürel E., Kaya Z. (2000): Doubled haploid plant production from unpollinated ovules of sugar beet ( Beta vulgaris L.). Plant Cell Reports, 19, 1155-1159  https://doi.org/10.1007/s002990000248
Hofinger B.J., Ankele E., Gülly Ch., Heberle-Bors E., Pfosser M.F. (2000): The involvement of the plastid genome in albino plant regeneration from microspores in wheat. In: Biotechnological Approaches for Utilization of Gametic Cells, COST 824 Final Meeting, Bled, July 1–5, 2000: 215–228.
Huang B. (1996): Gametoclonal variation in crop improvement. In: Jain S.M., Sopory S.K., Veilleux R.E. (eds): In vitro Haploid Production in Higher Plants. Current Plant Science and Biotechnology in Agriculture. Dordrecht, Springer: 73–91.
Inagaki M.N., Mujeeb-Kazi A. (1995): Comparison of polyhaploid production frequencies in crosses of hexaploid wheat with maize, pearl millet and sorghum. Breeding Science, 45: 157–161.
Inagaki M.N., Nagamine T., Mujeeb-Kazi A. (1997): Use of pollen storage and detached-tiller culture in wheat polyhaploid production through wide crosses. Cereal Research Communications, 25: 7–13.
Jacquard C. (2007): Pollen Embryogenesis in Barley (Hordeum vulgare L.): Importance of Pretreatment. [PhD. Thesis.] Reims, University of Reims Champagne-Ardenne. (in French)
Jahier J., Chèvre A.M., Eber F., Delourme R., Tanguy A.M. (1992): Plant Cytogenetic Techniques. INRA. (in French)
Jauhar P.P. (2003): Haploid and doubled haploid production in durum wheat by wide hybridization. In: Maluszynski M., Kasha K.J., Forster B.P., Szarejko I., (eds): Doubled Haploid Production in Crop Plants. A Manual. Netherlands, Kluwer Academic Publishers: 161–166.
Laurie D. A., Bennett M. D. (1988): The production of haploid wheat plants from wheat x maize crosses. Theoretical and Applied Genetics, 76, 393-397  https://doi.org/10.1007/BF00265339
Lu R., Chen Z., Gao R., He T., Wang Y., Xu H., Guo G., Li Y., Liu C., Huang J. (2016): Genotypes-independent optimization of nitrogen supply for isolated microspore cultures in barley. BioMed Research International, 2016: 1801646.
Makowska Katarzyna, Oleszczuk Sylwia (2014): Albinism in barley androgenesis. Plant Cell Reports, 33, 385-392  https://doi.org/10.1007/s00299-013-1543-x
Makowska K., Oleszczuk S., Zimny J. (2017): The effect of copper on plant regeneration in barley microspore culture. Czech Journal of Genetics and Plant Breeding, 53, 17-22  https://doi.org/10.17221/82/2016-CJGPB
Mdarhri-Alaoui Mériem, Saldi Najia, Chlyah Averil, Chlyah Hassan (1998): Obtention par gynogenèse in vitro de plantes haploïdes chlorophyliennes chez le blé dur. Comptes Rendus de l'Académie des Sciences - Series III - Sciences de la Vie, 321, 25-30  https://doi.org/10.1016/S0764-4469(97)89622-9
Mishra V.K., Goswami R. (2014): Haploid production in higher plant. International Journal of Chemical and Biological Sciences, 1: 25–45.
Murashige Toshio, Skoog Folke (1962): A Revised Medium for Rapid Growth and Bio Assays with Tobacco Tissue Cultures. Physiologia Plantarum, 15, 473-497  https://doi.org/10.1111/j.1399-3054.1962.tb08052.x
Niu Zhixia, Jiang Aixiang, Abu Hammad Wesam, Oladzadabbasabadi Atena, Xu Steven S., Mergoum Mohamed, Elias Elias M., Hartl L. (2014): Review of doubled haploid production in durum and common wheat through wheat × maize hybridization. Plant Breeding, 133, 313-320  https://doi.org/10.1111/pbr.12162
Picard E., Crambers E., Mihamou-Ziyyat A. (1994): Haplodiploidization: a multi-purpose tool for genetics and cereal breeding. In: What Future for Plant Breeding? Paris, AUPELF-UREF, John Libbey Eurotext: 355–369. (in French)
Khound Rituraj, Santra Meenakshi, Baenziger P. Stephen, Santra Dipak K. (2013): Effect of Cold-Mediated Pretreatment on Microspore Culture in Winter and Spring Wheat. American Journal of Plant Sciences, 04, 2259-2264  https://doi.org/10.4236/ajps.2013.411278
Slama-Ayed Olfa, Slim-Amara Hajer (2007): Production of doubled haploids in durum wheat (Triticum durum Desf.) through culture of unpollinated ovaries. Plant Cell, Tissue and Organ Culture, 91, 125-133  https://doi.org/10.1007/s11240-007-9281-9
Slama-Ayed O., Trifa Y., Slim-Amara H., De Buyser J., Picard E. (2010): Production of doubled haploids in Tunisian durum wheat (Triticum durum Desf.) cultivars through unpollinated ovary culture. Plant Mutation Reports, 2: 33–39.
Sibi M., Fakiri M. (1994): Gynogenesis in Moroccan genotypes barley (Hordeum vulgare). In: What Future for Plant Breeding? Paris, AUPELF-UREF, John Libbey Eurotext: 337–344. (in French)
Sibi M.L., Kobaissi A., Shekafandeh A. (2001): Green haploid plants from unpollinated ovary culture in tetraploid wheat (Triticum durum Defs.). Euphytica, 122: 351–359. https://doi.org/10.1023/A:1012991325228
Soriano M., Cistué L., Castillo A. M. (2008): Enhanced induction of microspore embryogenesis after n-butanol treatment in wheat (Triticum aestivum L.) anther culture. Plant Cell Reports, 27, 805-811  https://doi.org/10.1007/s00299-007-0500-y
Sriskandarajah Sridevy, Sameri Mohammad, Lerceteau-Köhler Estelle, Westerbergh Anna (2015): Increased Recovery of Green Doubled Haploid Plants from Barley Anther Culture. Crop Science, 55, 2806-  https://doi.org/10.2135/cropsci2015.04.0245
Ushiyama Tomohiko, Kuwabara Tatsuo, Yoshida Tomohiko (2015): Effects of Various Phytohormones On Haploid Wheat Production in Wheat x Maize Crosses. Plant Production Science, 10, 36-41  https://doi.org/10.1626/pps.10.36
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