The effects of sugars and growth regulators on embryo- and callusogenesis in isolated ovules culture of beetroot, Beta vulgaris L.

Document Type : Research Paper

Authors

Russian State Agrarian University - Moscow Timiryazev Agricultural Academy, Timiryazevskaya St. 49, Moscow

Abstract

Haploid techniques have risen interest among researchers and breeders as they significantly reduce the time of creating pure lines in breeding programs, especially for biennial vegetable crops. The only way to produce doubled haploids used in representatives of the genus Beta is in vitro culture of non-pollinated ovules. This is a rather laborious method that requires manual labor to isolate beet ovules from flower buds, followed by their inoculation onto a culture medium. This method also has its drawback - the development of clones from the somatic tissues surrounding the embryo sac. At the same time, the yield of embryoids and subsequent regenerations is on average 12-14% of isolated ovules introduced into in vitro culture in the most responsive genotypes of sugar beet and 8% in beetroot. The selection of the optimal cultivation conditions for each genotype makes it possible to maximize the yield of regenerant plants in the culture of isolated ovules. This research is devoted to study the effect of various types of carbohydrates and growth regulators in the culture media on embryogenesis and callusogenesis in isolated ovules culture of five beetroot genotypes. We obtained embryoids and callus in isolated ovules culture of all studied genotypes using sucrose-based culture medium, while on a glucose- and fructose-based culture media or their combination, the number of callus and embryoids was much lower. Additionally, it has been shown that glucose has more negative effect on embryo- and callusogenesis than fructose. The addition of mannitol to the culture medium had a stimulatory effect on the ovules for one genotype only. The study of various combinations of phytohormones on 5 beetroot genotypes showed that gibberellic acid reduces the embryogenesis response of ovules in all genotypes.

Keywords


Baranski, R 1996, In vitro gynogenesis in beetroot (Beta vulgaris L.): effects of ovule culture conditions. Acta Societatis Botanicorum Poloniae, 65: 57-60.
D’Halluin, K & Keimer, B 1986, Production of haploid sugar beets (Beta vulgaris L.) by ovule culture. Genetic Manipulation in Plant Breeding. Berlin. DOI:10.1515/9783110871944.
De La Fuente, GN, Frei, UK & Lübberstedt, T 2013, Accelerating plant breeding. Trends Plant Science, 18: 667-672. DOI:10.1016/j.tplants.2013.09.001.
Doctrinal, M, Sangwan, RS & Sangwan-Norreel, BS 1989, In vitro gynogenesis in Beta vulgaris L.: Effects of plant growth regulators, temperature, genotypes and season. Plant Cell, Tissue and Organ Culture, 17: 1-12. DOI:10.1007/BF00042276
Gurel, S, Gurel, E & Kaya, Z 2000, Doubled haploid plant production from unpollinated ovules of sugar beet (Beta vulgaris L.). Plant Cell Reports, 19:1155-1159. DOI:10.1007/s002990000248
Klimek-Chodacka, M & Baranski, R 2013, Comparison of haploid and doubled haploid sugar beet clones in their ability to micropropagate and regenerate. Electronic Journal of Biotechnology, 16: 1, DOI:10.2225/vol16-issue2- fulltext-3
Lux, H, Herrmann, NL & Wetzel, C 1990, Production on haploid sugarbeet (Beta vulgaris L.) by culturing unpollinated ovules. Plant Breed, 104: 177-183. DOI:10.1111/j.1439-0523.1990.tb00420.x
Pazuki, A, Aflaki, F, Gürel, E et al. 2018, Gynogenesis Induction in Sugar Beet (Beta vulgaris) Improved by 6-Benzylaminopurine (BAP) and Synergized with Cold Pretreatment. Sugar Tech, 20: 69-77. DOI:10.1007/s12355-017-0522-x
Seman, I & Farago J 1990, In vitro cultivation of unfertilized ovules of sugar beet. Embryology and seed reproduction: XI International  Symposium, Leningrad, USSR, July 3-4, Leningrad. 412.39
Śliwińska, E, Łukaszewska, E 2005, Polysomaty in growing in vitro sugar-beet (Beta vulgaris L.) seedlings of different ploidy level. Plant Science, 168: 1067–1074. DOI:10.1016/j.plantsci.2004.12.003
Tomaszewska-Sowa, M 2010, Cytometric analyses of sugar beet (Beta vulgaris L.) plants regenerated from unfertilized ovules cultured in vitro. Plant Breed, 2: 231-235.
Van Geyt, J, Speckmann, GJ, D'Halluin, K et al. 1987, In vitro induction of haploid plants from unpollinated ovules and ovaries of the sugarbeet (Beta vulgaris L.). Theoretical and Applied Genetics, 73: 920-925. DOI:10.1007/BF00289399
Vasilchenko, EN, Zhuzhzhalova, TP, Vashchenko, TG, Zemlianukhina, OA, Karpechenko, NA & Podvigina, OA 2017, Features of the formation of haploid regenerants of sugar beet in culture in vitro. Bulletin of Voronezh State Agrarian University, 3: 57-66.
Wremerth Weich, E & Levall, MW 2003, Doubled haploid production of sugar beet (Beta vulgaris L.): Published protocols for other crop plant species. In Doubled Haploid Production in Crop Plants: A Manual. Kluwer Academic Publishers. DOI:10.1007/978-94-017-1293-4_38
Zhuzhzhalova, TP, Kolesnikova, EO, Vasilchenko, EN & Cherkasova, NN 2020, Biotechnological methods as a tool for efficient sugar beet breeding. Vavilov Journal of Genetics and Breeding, 24: 40-47. DOI:10.18699/VJ20.593