Abstract
The present study reports an efficient in vitro plant regeneration system for amphidiploid (2n = 42) spelt wheat (Triticum spelta L.). Plant regeneration from mature embryos of winter spelt variety “Europe” was carried out as a two-step process. The first step was callus induction on a medium supplemented with 2 mg L−1 dichlorophenoxyacetic acid and 10 mg L−1 silver nitrate resulting in 96% callus formation. The second step resulted in 30% plant regeneration frequency from calluses transferred to Murashige and Skoog medium supplemented with 2 mg L−1 6-benzylaminopurine, 0.5 mg L−1 1-naphthaleneacetic acid, and 10 mg L−1 silver nitrate. Regeneration medium supplemented with 1 mg L−1 6-benzylaminopurine, 0.2 mg L−1 picloram, and the hormone-free medium turned out to be less effective in our experiments. Regenerated plants formed roots after transfer to half-strength Murashige and Skoog medium supplemented with 0.7 mg L−1 indole-3-butyric acid. About one-third of the resulting regenerated plants that formed roots were transferred to soil. The inter-simple sequence repeat markers were used to confirm the genetic homogeneity of regenerated plants. Thus, our regeneration protocol can be used for in vitro regeneration of spelt plants from mature embryos with minimal risk of genetic variability and provides an essential step towards developing an efficient strategy for spelt genetic transformation.
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Abid N, Maqbool A, Malik KA (2014) Screening commercial wheat (Triticum aestivum L.) varieties for Agrobacterium mediated transformation ability. Pak J Agr Sci 51:83–89
Abou-Deif MH, Rashed MA, Sallam MAA, Mostafa EAH, Ramadan WA (2013) Characterization of twenty wheat varieties by ISSR markers. Middle-East J Sci Res 15:168–175
Adero MO, Syombua ED, Asande LK, Amugune NO, Mulanda ES, Macharia G (2019) Somatic embryogenesis and regeneration of Kenyan wheat (Triticum aestivum L.) genotypes from mature embryo explants. Afr J Biotechnol 18:689–694
Ahmad A, Zhong H, Wang W, Sticklen MB (2002) Shoot apical meristem: In vitro regeneration and morphogenesis in wheat (Triticum aestivum L.). In Vitro Cell Dev Biol-Plant 38:163–167
Baday SJS (2018) In vitro study of the callus induction of two varieties of wheat seeds by plant growth regulators. Biochem Cell Arch 18:2067–2071
Bommineni VR, Jauhar PP (1996) Regeneration of plantlets through isolated scutellum culture of durum wheat. Plant Sci 116:197–203
Bouiamrine EH, Diouri M, El-Halimi R (2012) Assessment of somaclonal variation in regenerated plants from immature embryos culture of durum wheat. Int J Agric Biol 14:941–946
Chen THH, Lazar MD, Scoles GJ, Gusta LV, Kartha KK (1987) Somaclonal variation in a population of winter wheat. J Plant Physiol 130:27–36
Dargahlou SA, Uliaie ED, Bandehagh A (2017) Callus induction and plant regeneration from mature embryos of some Iranian wheat (Triticum aestivum L.) genotypes. J Bio Env Sci 10:275–283
Doyle JJ, Doyle JL (1990) Isolation of plant DNA from fresh tissue. Focus 12:13–15
Fujii T (1970) Callus formation in wheat anthers. Wheat Inform Serv, Kyoto Univ 31:1–2
Gomez-Becerra HF, Erdem H, Yazici A, Tutus Y, Torun B, Ozturk L, Cakmak I (2010) Grain concentrations of protein and mineral nutrients in a large collection of spelt wheat grown under different environments. J Cereal Sci 52:342–349
Guzman C, Medina-Larque AS, Velu G, Gonzalez-Santoyo H, Singh RP, Huerta-Espino J, Ortiz-Monasterio I, Pena RJ (2014) Use of wheat genetic resources to develop biofortified wheat with enhanced grain zinc and iron concentrations and desirable processing quality. J Cereal Sci 60:617–622
Hu T, Metz S, Chay C, Zhou HP, Biest N, Chen G, Cheng M, Feng X, Radionenko M, Lu F, Fry J (2003) Agrobacterium-mediated large-scale transformation of wheat (Triticum aestivum L.) using glyphosate selection. Plant Cell Rep 21:1010–1019
Ihsan SM, Jabeen M, Ilahi I (2003) In vitro callus induction, its proliferation and regeneration in seed explants of wheat (Triticum aestivum L.). Pak J Bot 35:209–217
Kacem NS, Muhovski Y, Djekoun A, Watillon B (2017) Molecular characterization of genetic variation in somaclones of durum wheat (Triticum durum Desf) using SSR markers. Eur Sci J 13:426–437
Karadimova M (1993) Somaclonal variation in wheat. Biotechnol Biotechnol Eq 7:26–29
Kumar R, Mamrutha HM, Kaur A, Venkatesh K, Grewal A, Kumar R, Tiwari V (2017) Development of an efficient and reproducible regeneration system in wheat (Triticum aestivum L.). Physiol Mol Biol Plants 23:945–954
Kyriienko AV, Parii MF, Kuchuk MV, Symonenko YV, Shcherbak NL (2019) Development of an effective technique for callus formation from mature embryos of Triticum spelta L. and Triticum aestivum L. Plant Varieties Studying Protect 15:259–265
Laemmli UK (1970) Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227:680–685
Lantos C, Bóna L, Nagy E, Békés F, Pauk J (2018) Induction of in vitro androgenesis in anther and isolated microspore culture of different spelt wheat (Triticum spelta L.) genotypes. Plant Cell Tiss Org Cult 133:385–393
Liubych VV, Hospodarenko HM, Poltoretskyi SP (2017) Quality features of spelt wheat grain. LAP LAMBERT Academic Publishing, Saarbrücken
Mathias RJ, Simpson ES (1986) The interaction of genotype and culture medium on the tissue culture responses of wheat (Triticum aestivum L. em. thell) callus. Plant Cell Tiss Org Cult 7:31–37
Murashige T, Skoog F (1962) A revised medium for rapid growth and bio assays with tobacco tissue cultures. Physiol Plant 15:473–497
Nasircilar AG, Turgut K, Fiskin K (2006) Callus induction and plant regeneration from mature embryos of different wheat genotypes. Pak J Bot 38:637–645
Nizamani F, Nizamani GS, Nizamani MR, Ahmed S, Ahmed N (2016) Callus induction and plant regeneration from immature embryos culture in spring wheat (Triticum aestivum L.) varieties. Int J Biol Res 1:28–32
Özgen M, Türet M, Altınok S, Sancak C (1999) Efficient callus induction and plant regeneration from mature embryo culture of winter wheat (Triticum aestivum L.) genotypes. Plant Cell Rep 18:331–335
Ozias-Akins P, Vasil IK (1982) Plant regeneration from cultured immature embryos and inflorescences Triticum aestivum L. (wheat): evidence for somatic embryogenesis. Protoplasma 110:95–105
Parmar SS, Sainger M, Chaudhary D, Jaiwal PK (2012) Plant regeneration from mature embryo of commercial Indian bread wheat (Triticum aestivum L.) cultivars. Physiol Mol Biol Plants 18:177–183
Porebski S, Bailey LG, Baum BR (1997) Modification of a CTAB DNA extraction protocol for plants containing high polysaccharide and polyphenol components. Plant Mol Biol Rep 15:8–15
Przetakiewicz A, Orczyk W, Nadolska-Orczyk A (2003) The effect of auxin on plant regeneration of wheat, barley and triticale. Plant Cell Tiss Org Cult 73:245–256
Schmid J (1990) In vitro production of haploids in Triticum spelta. In: Bajaj YPS (ed) Wheat, Biotechnology in agriculture and forestry, Vol, vol 13. Springer, Berlin, Heidelberg, pp 363–381
Shah MI, Jabeen M, Ilahi I (2003) In vitro callus induction, its proliferation and regeneration in seed explants of wheat (Triticum aestivum L.) var.lu-26s. Pak J Bot 35:209–217
Songstad DD, Duncan DR, Widholm JM (1988) Effect of l-aminocyclopropane-l-carboxylic acid, silver nitrate, and norbornadiene on plant regeneration from maize callus cultures. Plant Cell Rep 7:262–265
Symillides Y, Henry Y, De Buyser J (1995) Analysis of Chinese Spring regenerants obtained from short- and long-term wheat somatic embryogenesis. Euphytica 82:263–268
Upadhyaya G, Sen M, Roy A (2015) In vitro callus induction and plant regeneration of rice (Oryza sativa L.) var. ‘Sita’, ‘Rupali’ and ‘Swarna Masuri’. Asian J Plant Sci Res 5:24–27
Vu NT, Chin J, Pasco JA, Kovács A, Wing LW, Békés F, Suter DAI (2015) The prevalence of wheat and spelt sensitivity in a randomly selected Australian population. Cereal Res Commun 43:97–107
Wu LM, Wei YM, Zheng YL (2006) Effects of silver nitrate on the tissue culture of immature wheat embryos. Russ J Plant Physiol 53:530–534
Xhulaj DB (2018) Effect of plant growth regulators on in vitro regeneration from mature wheat (Triticum aestivum L.) embryo explants. Agrotechnology 7:112–113
Yasmin S, Khan IA, Khatri A, Seema N, Nizamani GS, Arain MA (2009) In vitro plant regeneration in bread wheat (Triticum аestivum L.). Pak J Bot 41:2869–2876
Acknowledgements
The seed material was provided by LLC “Ukrainian Scientific Institute of Breeding” (Ukraine). The work was performed within the fundamental departmental topics of the Department of Genetic Engineering Institute of Cell Biology and Genetic Engineering of NAS of Ukraine: III-8-17 “Study of the functioning of heterologous genes and their impact on the adaptive characteristics of plant systems under biotic and abiotic stresses” (№ state registration 0117U002589, 2017–2021).
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Editor: Tom Clemente
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Kyriienko, A., Shcherbak, N.L., Kuchuk, M.V. et al. In vitro plant regeneration from mature embryos of amphidiploid spelt Triticum spelta L.. In Vitro Cell.Dev.Biol.-Plant 57, 856–863 (2021). https://doi.org/10.1007/s11627-021-10158-4
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DOI: https://doi.org/10.1007/s11627-021-10158-4