FAST DIRECT REGENERATION OF PLANTS FROM NODAL EXPLANTS OF Stevia rebaudiana Bert.
Romuald DolińskiInsitute of Plant Genetics, Breeding and Biotechnology, University of Life Sciences in Lublin, Akademicka 15, 20-950 Lublin, Poland
Krzysztof KowalczykInsitute of Plant Genetics, Breeding and Biotechnology, University of Life Sciences in Lublin, Akademicka 15, 20-950 Lublin, Poland
In the preceding research, stevia has been typically cloned in vitro using two media, on which the shoots were formed (3–6 weeks), and on the other they were rooted (3–5 weeks). This study aimed at finding the possibility for rapid stevia propagation from large nodal explants using the MS basal medium [Murashige and Skoog 1962], with low auxin concentrations (0.5, 1 and 2 mg‧dm–3). The plants were obtained as soon as after three weeks. The best results were obtained from media with various concentrations of the indole-3-acetic acid (IAA) and the highest concentration of phenylacetic acid (PAA). Plants were formed by 83.9-86.0% of explants, they had high weight (234−253 mg), two shoots measuring 2.07−2.37 cm and 5.8−8.3 roots measuring 1.00–1.24 cm. Mean plant weight was lowest on the media with indole-3-butyric acid (IBA) (185–192 mg). Both explant buds formed single shoots, but their development was typically uneven. The differences in the length and weight of shoots were lowest on the media with IAA and at lower PAA concentrations. Plants from the media with IAA and the control medium were distinguished by a higher number of nodes. The percentage share of shoots in the total plant weight was highest on the media with PAA (62.1–62.7%), and lowest at higher concentrations of α-naphthaleneacetic acid (NAA) (47.9 and 48.9%). Parts of explants immersed in media developed callus, and the highest amounts of this tissue were found in the media with NAA. 92.3% of plants survived the acclimatization. The applied procedure may be used for rapid in vitro cloning of selected stevia genotypes. The use of one medium enables reduction of seedling production costs. Moreover, cyclical cloning and extending the production scale is possible.
Keywords:one-step micropropagation, nodal explants, low concentrations auxins
Ahmed, M.B., Salahin, M., Karim, R., Razvy, M. A., Hannan, M.M., Sultana, R., Hossain, M., Islam, R. (2007). An efficient method for in vitro clonal propagation of a newly introduced sweetener plant (Stevia rebaudiana Bertoni) in Bangladesh. Am.-Euras. J. Sci. Res., 2(2), 121–125.
Anbazhagan, M., Kalpana, M., Rajendran, R., Natarajan, V., Dhanavel, D. (2010). In vitro production of Stevia rebaudiana Bertoni. Emir. J. Food Agric., 22(3), 216–222.
Brandle, J.E., Starratt, A.N., Gijzen, M. (1998). Stevia rebaudiana: its agricultural, biological and chemical properties. Can. J. Plant Sci., 78, 527–536.
Carneiro, J.W.P., Muniz, A.S., Guedes, T.A. (1997). Greenhouse bedding plant production of Stevia rebaudiana (Bert) bertoni. Can. J. Plant Sci., 77, 473–474.
Das, A., Gantait, S., Mandal, N. (2011). Micropropagation of an elite medicinal plant: Stevia rebaudiana Bert. Int. J. Agric. Res., 6(1), 40–48.
Driver, J.A., Kuniyuki, A.H. (1984). In vitro propagation of Paradox walnut rootstock. Hort. Sci., 19, 507–509.
Genus, J.M., Buyse, J., Vankeirsbilck, A., Temme, E.H. (2007). Metabolism of stevioside by healthy subjects. Exp. Biol. Med., 232, 163–173.
Grenby, T.H. (1991). Update on low-calorie sweeteners to benefit dental health. Int. J. Dent., 41, 217–224.
Huxtable, R.J. (2002). Pharmacology and toxicology of stevioside, rebaudioside A, and steviol. In: Stevia, the genus Stevia. Medicinal and aromatic plants − industrial profiles, Kinghorn A. D. (Ed.). Vol. 19. Tylor Francis Group, London−New York, 160–178.
Khalil, S.A., Zamir, R., Ahmed, N. (2014). Selection of suitable propagation method for consistent plantlets production in Stevia rebaudiana (Bertoni). Saudi J. Biol. Sci., 21, 566–573.
Lata, H., Chandra, S., Wang, Y.H., Raman, V., Khan, I.A. (2013). TDZ-induced high frequency plant regeneration through direct shoot organogenesis in Stevia rebaudiana Bertoni: an important medicinal plant and a natural sweetener. Am. J. Plant Sci., 4, 117–128.
Lemus-Mondaca, R., Vega-Galez, A., Zura-Bravo, L., Ah-Hen, K. (2012). Stevia rebaudiana Bertoni, source of a high-potency natural sweetener: A comprehensive review on the biochemical, nutritional and functional aspects. Food Chem., 132, 1121–1132.
Lloyd, G., McCown, B. (1980). Commercially-feasible micropropagation of mountain laurel, Kalmia latifolia, by use of shot-tip culture. Inter Plant Prop. Soc. Proc., 30, 421–445.
Mathur, M., Begum, T. (2015). Shootles regeneration and tissue culture studies on Stevia rebaudiana Bertoni and Terminalia bellerica Roxb. Int. J. Rec. Biotech., 3(1), 25–35.
Murashige, T., Skoog, F. (1962). A revised medium for rapid growth and bioassays with tobacco tissue cultures. Physiol. Plant., 15, 473–497.
Peixe, A., Calado, J., Carlos, R. (2015). A one-step procedure for in vitro micropropagation of stevia (Stevia rebaudiana Bertoni). Acta Hortic., 1083, 295–301.
Rafiq, M., Dahot, M.U., Mangrio, S.M., Naqvi, H.A., Qarshi, I.A. (2007). In vitro clonal propagation and biochemical analysis of field established Stevia rebaudiana Bertoni. Pak. J. Bot., 39(7), 2467–2474.
Ramesh, K., Singh, V., Megeji, N.W. (2006). Cultivation of Stevia [Stevia rebaudiana (Bert.) Bertoni]: a comprehensive review. Adv. Agron., 89, 137–177.
Razak, U.N.A.A., Ong, C.B., Yu, T.S., Lau, L.K. (2014). In vitro micropropagation of Stevia rebaudiana Bertoni in Malaysia. Braz. Arch. Biol. Technol., 57(1), 23–28.
Rosales, C., Brenes, J., Salas, K., Arce-Solano, S., Abdelnour-Esquivel, A. (2018). Micropropagation of Stevia rebaudiana in the temporary immersion systems as an alternative horticultural production method. Rev. Chapingo Ser. Horticultura 24(1), 69–84.
Savita, S.M., Shella, K., Sunanda, S., Shankar, A.G., Ramakrishna, P. (2004). Stevia rebaudiana – a functional component for food industry. J. Human Ecol. 15, 261–264.
Soejarto, D.D. (2002). Botany of Stevia rebaudiana. In: Stevia, the genus Stevia. Medicinal and aromatic plants − industrial profiles, Kinghorn A. D. (Ed.). Vol. 19. Tylor Francis Group, London−New York, 18−39.
Taleje, N., Hamidoghli, S., Hamidoghli, Y. (2012). In vitro plantlet propagation of Stevia rebaudiana Bertoni. South West. J. Hortic. Biol. Environ., 3(1), 99–108.
Tateo, F., Mariotti, M., Bononi, M., Lubian, E., Martello, S., Cornara, L. (1998). Stevioside content and morphological variability in a population of Stevia rebaudiana (Bertoni) from Paraguay. Ital. J. Food. Sci. 10(3), 261–267.
Verma, S., Yadav, K., Singh, N. (2011). Optimization of the protocols for surface sterilization, regeneration and acclimatization of Stevia rebaudiana Bertoni. Am.-Eur. J. Agric. Environ. Sci., 11(2), 221–227.
Yücesan, B., Büyükgöçmen, R., Mohammed, A., Sameeullah, M., Altuğ, C., Gürel, S., Gürel, E. (2016). An efficient regeneration system and steviol glycoside analysis of Stevia rebaudiana Bertoni, a source of natural high-intensity sweetener. In Vitro Cell. Dev. Biol.-Plant, 52, 330–337.
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