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Production of Doubled Haploid Plants in Cucumber (Cucumis sativus L.) Through Anther Culture

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Doubled Haploid Technology

Part of the book series: Methods in Molecular Biology ((MIMB,volume 2289))

Abstract

As in any other economically important crop, the possibility of producing fully homozygous, doubled haploid lines in cucumber allows for faster and cheaper breeding. At present, the fastest way to doubled haploidy is the production of cucumber haploid plants and duplication of their chromosomes to make them doubled haploid. In this chapter, we describe a complete protocol to successfully produce cucumber doubled haploid plants, including the evaluation of their ploidy level by flow cytometry. Briefly, this protocol involves a first step of anther culture to induce microspores to divide and proliferate forming calli. The calli produced are isolated from anthers and transferred first to a liquid medium and then to a solid medium to induce organogenesis. Organogenic shoots will eventually give rise to entire DH plants.

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References

  1. FAOSTAT (2020). http://www.fao.org/faostat. Last checked March 2020

  2. Gałązka J, Niemirowicz-Szczytt K (2013) Review of research on haploid production in cucumber and other cucurbits. Folia Hortic 25:67–78. https://doi.org/10.2478/fhort-2013-0008

    Article  Google Scholar 

  3. Dong Y-Q, Zhao W-X, Li X-H, Liu X-C, Gao N-N, Huang J-H, Wang W-Y, Xu X-L, Tang Z-H (2016) Androgenesis, gynogenesis, and parthenogenesis haploids in cucurbit species. Plant Cell Rep 35:1991–2019. https://doi.org/10.1007/s00299-016-2018-7

    Article  CAS  PubMed  Google Scholar 

  4. Claveria E, Garcia-Mas J, Dolcet-Sanjuan R (2005) Optimization of cucumber doubled haploid line production using in vitro rescue of in vivo induced parthenogenic embryos. J Am Soc Hortic Sci 130(4):555–560

    Article  Google Scholar 

  5. Ebrahimzadeh H, Soltanloo H, Shariatpanahi ME, Eskandari A, Ramezanpour SS (2018) Improved chromosome doubling of parthenogenetic haploid plants of cucumber (Cucumis sativus L.) using colchicine, trifluralin, and oryzalin. Plant Cell Tissue Organ Cult 135(3):407–417. https://doi.org/10.1007/s11240-018-1473-y

    Article  CAS  Google Scholar 

  6. Truong-Andre I (1988) In vitro haploid plants derived from pollination by irradiated pollen on cucumber. In: Eucarpia meeting on cucurbit genetics and breeding, Montfavet (France), 31 May-2 Jun 1988. INRA

    Google Scholar 

  7. Çaglar G, Abak K (1999) Progress in the production of haploid embryos, plants and doubled haploids in cucumber (C. Sativus L.) by gamma irradiated pollen, in Turkey. Acta Hortic 492:317–322

    Article  Google Scholar 

  8. Dolcet-Sanjuan R, Claveria E, Garcia-Mas J (2004) Cucumber (Cucumis sativus L.) dihaploid line production using in vitro rescue of in vivo induced parthenogenic embryos. Acta Hortic 725:837–844

    Google Scholar 

  9. Tantasawat PA, Sorntip A, Pornbungkerd P (2015) Effects of exogenous application of plant growth regulators on growth, yield, and in vitro Gynogenesis in cucumber. Hortscience 50(3):374–382. https://doi.org/10.21273/hortsci.50.3.374

    Article  CAS  Google Scholar 

  10. Diao W-P, Jia Y-Y, Song H, Zhang X-Q, Lou Q-F, Chen J-F (2009) Efficient embryo induction in cucumber ovary culture and homozygous identification of the regenetants using SSR markers. Sci Hortic 119(3):246–251. https://doi.org/10.1016/j.scienta.2008.08.016

    Article  CAS  Google Scholar 

  11. Gémes-Juhász A, Balogh P, Ferenczy A, Kristóf Z (2002) Effect of optimal stage of female gametophyte and heat treatment on in vitro gynogenesis induction in cucumber (Cucumis sativus L.). Plant Cell Rep 21(2):105–111. https://doi.org/10.1007/s00299-002-0482-8

    Article  CAS  Google Scholar 

  12. Plapung P, Khamsukdee S, Potapohn N, Smitamana P (2014) Screening for cucumber mosaic resistant lines from the ovule culture derived double haploid cucumbers. Am J Agric Biol Sci 9(3):261–269

    Article  Google Scholar 

  13. Ashok Kumar HG, Murthy HN (2004) Effect of sugars and amino acids on androgenesis of Cucumis sativus. Plant Cell Tissue Organ Cult 78(3):201–208. https://doi.org/10.1023/b:ticu.0000025637.56693.68

    Article  Google Scholar 

  14. Song H, Lou QF, Luo XD, Wolukau JN, Diao WP, Qian CT, Chen JF (2007) Regeneration of doubled haploid plants by androgenesis of cucumber (Cucumis sativus L.). Plant Cell Tissue Organ Cult 90(3):245–254. https://doi.org/10.1007/s11240-007-9263-y

    Article  CAS  Google Scholar 

  15. Hamidvand Y, Abdollahi MR, Chaichi M, Moosavi SS (2013) The effect of plant growth regulators on callogenesis and gametic embryogenesis from anther culture of cucumber (Cucumis sativus L.). Int J Agric Crop Sci 5(10):1089

    Google Scholar 

  16. Abdollahi MR, Najafi S, Sarikhani H, Moosavi SS (2016) Induction and development of anther-derived gametic embryos in cucumber (Cucumis sativus L.) by optimizing the macronutrient and agar concentrations in culture medium. Turk J Biol 40(3):571–579

    Article  CAS  Google Scholar 

  17. Xie M, Qin L-Y, Pan J-S, He H-L, Wu A-Z, Cai R (2005) Flower morphogenesis and microspore development versus anther culture of cucumber. Acta Bot Boreal-Occid Sin 25(6):1096

    Google Scholar 

  18. Zhan Y, J-f C, Malik AA (2009) Embryoid induction and plant regeneration of cucumber (Cucumis sativus L.) through microspore culture. Acta Hortic Sin 36(2):221–226

    CAS  Google Scholar 

  19. Asadi A, Zebarjadi A, Abdollahi MR, Seguí-Simarro JM (2018) Assessment of different anther culture approaches to produce doubled haploids in cucumber (Cucumis sativus L.). Euphytica 214(11):216. https://doi.org/10.1007/s10681-018-2297-x

    Article  CAS  Google Scholar 

  20. Custers J (2003) Microspore culture in rapeseed (Brassica napus L.). In: Maluszynski M, Kasha KJ, Forster BP, Szarejko I (eds) Doubled haploid production in crop plants. Kluwer Academic Publishers, Dordrecht, The Netherlands, pp 185–193

    Chapter  Google Scholar 

  21. Malepszy S, Niemirowicz-Szczytt K (1991) Sex determination in cucumber (Cucumis sativus) as a model system for molecular biology. Plant Sci 80(1–2):39–47

    Article  Google Scholar 

  22. Liberatore CM, Calabuig-Serna A, Rodolfi M, Chiancone B, Seguí-Simarro JM (2019) Phenological phases of flowering in hop (Humulus lupulus L.) and their correspondence with microsporogenesis and microgametogenesis. Sci Hortic 256:108639. https://doi.org/10.1016/j.scienta.2019.108639

    Article  Google Scholar 

  23. Seguí-Simarro JM, Nuez F (2005) Meiotic metaphase I to telophase II is the most responsive stage of microspore development for induction of androgenesis in tomato (Solanum lycopersicum). Acta Physiol Plant 27(4b):675–685

    Article  Google Scholar 

  24. Parra-Vega V, González-García B, Seguí-Simarro JM (2013) Morphological markers to correlate bud and anther development with microsporogenesis and microgametogenesis in pepper (Capsicum annuum L.). Acta Physiol Plant 35(2):627–633. https://doi.org/10.1007/s11738-012-1104-x

    Article  Google Scholar 

  25. Salas P, Rivas-Sendra A, Prohens J, Seguí-Simarro JM (2012) Influence of the stage for anther excision and heterostyly in embryogenesis induction from eggplant anther cultures. Euphytica 184(2):235–250. https://doi.org/10.1007/s10681-011-0569-9

    Article  Google Scholar 

  26. Seguí-Simarro JM, Nuez F (2008) Pathways to doubled haploidy: chromosome doubling during androgenesis. Cytogenet Genome Res 120(3–4):358–369. https://doi.org/10.1159/000121085

    Article  PubMed  Google Scholar 

  27. Kurtar ES (2018) The effects of anti-mitotic agents on dihaploidization and fertility in winter squash (Cucurbita maxima Duch.) and pumpkin (Cucurbita moschata Duch.) androgenic haploids. Acta Sci Pol-Hortorum Cultus 17(5):3–14. https://doi.org/10.24326/asphc.2018.5.1

    Article  Google Scholar 

  28. Murashige T, Skoog F (1962) A revised medium for rapid growth and bioassays with tobacco tissue cultures. Physiol Plant 15:473–479

    Article  CAS  Google Scholar 

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Acknowledgments

This work was supported by grants AGL2017-88135-R and PID2020-115763RB-I00 to JMSS from Spanish MICINN jointly funded by FEDER.

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Correspondence to Jose M. Seguí-Simarro .

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Asadi, A., Seguí-Simarro, J.M. (2021). Production of Doubled Haploid Plants in Cucumber (Cucumis sativus L.) Through Anther Culture. In: Segui-Simarro, J.M. (eds) Doubled Haploid Technology. Methods in Molecular Biology, vol 2289. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-1331-3_4

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  • DOI: https://doi.org/10.1007/978-1-0716-1331-3_4

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  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-0716-1330-6

  • Online ISBN: 978-1-0716-1331-3

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