Callus Induction and Micropropagation of Lilium candidum L. Using Stem Bulbils and Confirmation of Genetic Stability via SSR-PCR

Year 2020, Volume: 7 Issue: 4, 286 - 296, 15.12.2020

Hilal Büşra Tokgöz , Filiz Altan

https://doi.org/10.21448/ijsm.753053

Cited By: 4

Abstract

Natural populations of Lilium candidum L. are remarkably affected by biotic and abiotic factors therefore there is a requirement to develop effective micropropagation protocol to provide mass production, multiplication and conservation of these plants. For this reason, this study was aimed to develop an efficient micropropagation method for multiple shoot production via somatic embryogenesis induced from L. candidum stem bulbils and also to determine the genetic stability of in vitro grown plants using SSR markers. The obtained results of this study are the first comprehensive reports including an investigation of genetic fidelity on somatic embryogenesis of L. candidum. After surface sterilization of bulbils, the calculated regeneration percentage of them was 89.5% and the callus induction was achieved using leaf segments of in vitro grown bulbils. The well formed somatic embryos were obtained from smooth whitish-yellow colored calli and these somatic embryos produced well formed healthy L. candidum seedlings with adventitious roots. All rooted seedlings were easily adapted to greenhouse conditions and the genetic stability of in vitro grown seedlings were determined by using SSR-PCR technique and it was calculated as 100%.

Keywords

Callus Culture, Genetic Stability, Lilium candidum L., Somatic Embryogenesis, SSR

Supporting Institution

The Scientific and Technological Resarch Council of Turkey (TUBİTAK)

Project Number

TOVAG- 217O009

Thanks

We thank to Assoc. Prof. Dr. Ergun KAYA for support and we also thank to Assoc. Prof. Dr. Hasan YILDIRIM and Assist Prof. Dr. Ademi PİRHAN for material support.

References

• Baytop, T. (1984). Türkiye’de Bitkiler ile Tedavi, İstanbul Üniversitesi, Yayın No: 3255, Eczacılık Fakültesi Yayın No: 40, İstanbul.
• Khan, N., Barba-Gonzalez, R., Lim, K.B., Ramanna, M.S., Visser, R.G.F., Van Tuyl, J.P. (2010). Relevance of Unilateral and Bilateral Sexual Polyploidisation in Relation to Intergenomic Recombination and Introgression in Lilium Species Hybrids. Euphytica 171, 157–173. https://doi.org/10.1007/s10681-009-9998-0
• Azadi, P., Kosh, K.I.U. (2007). Micropropagation of Lilium ledebori (Baker) as Effected by Plant Growth Regulators, Sucrose Concentrations, Harvesting Time and Cold Treatment. Electronic Journal of Biotechnology, 10(4), 587–591. https://doi.org/10.2225/vol10-issue4-fulltext-7
• Özüdoğru, E.A., Kaya, E., Kirdok, E., Issever-Ozturk, S. (2011). In vitro Propagation From Young and Mature Explants of Thyme (Thymus vulgaris and T. longicaulis) Resulting in Genetically Stable Shoots. In Vitro Cellular & Developmental Biology–Plant, 47(2), 309–320. https://doi.org/10.1007/s11627-011-9347-6
• Yeole, M.P., Gholse, Y.N., Gurunani, S.G., Dhole, S.M. (2016). Plant Tissue Culture Techniques: A Review for Future View. Critical Review in Pharmaceutıcal Sciences 5, 16–24.
• Klimaszewska, K., Park, Y.S., Bonga, J.M. (2011). Tissue Culture Research at The CFS: Its History, Current Status and Potential Benefits. Canadian Forest Service Publications, Catalog ID: 32295.
• Korban, S.S., Sul, I.W. (2007). Micropropagation of Coast Redwood (Sequoia Sempervirens). In: Jain SM, Häggman H (eds) Protocols for Micropropagation of Woody Trees and Fruits (pp. 23-32). Springer, Dordrecht.
• Yam, T.W., Arditti, J. (2018). Orchid Micropropagation: An Overview of Approaches and Methodologies. In: Lee YI, Yeung ET (eds) Orchid Propagation: From Laboratories to Greenhouses Methods and Protocols. Springer Protocols Handbooks. Humana Press, New York.
• Gaj, M.D. (2004). Factors Influencing Somatic Embryogenesis Induction and Plant Regeneration with Particular Reference to Arabidopsis thaliana (L.) Heynh. Plant Growth Regulation, 43, 27–47. https://doi.org/10.1023/B:GROW.0000038275.29262.fb
• Singh, N.D., Sahoo, L., Sarin, N.B., Jaiwal, P.K. (2003). The Effect of TDZ on Organogenesis and Somatic Emyogenesis in Pigeonpea (Cajanus cajan L. Millsp). Plant Science, 164, 341–347.
• Butiuc–Keul, A., Farkas, A., Cristea, V. (2016). Genetic Stability Assessment of in vitro Plants by Molecular Markers. Studia Universitatis Babeş-Bolyai Biologia, 61(1), 107-114.
• Kalia, R.K., Rai, M.K., Kalia, S., Singh, R., Dhawan, A.K. (2011). Microsatelite Markers: An Overview of The Recent Progress in Plants. Euphytica, 177, 309–334. https://doi.org/10.1007/s10681-010-0286-9
• Varshney, A., Lakshmikumaran, M., Srivastava, P.S., Dhawan, V. (2001). Establishment of Genetic Fidelity of in vitro-raised Lilium Bulblets Through RAPD Markers. In Vitro Cell Developmental Biology-Plant, 37(2), 227–231. https://doi.org/10.1007/s11627-001-0040-z
• Khawar, K.M., Cocu, S., Parmaksiz, I., Sarihan, E.O., Özcan, S. (2005). Mass proliferation of Madonna Lily (Lilium candidum L) Under in vitro Conditions. Pakistan Journal of Botany, 37(2), 243–248.
• Liu, X., Yang, G. (2012). Adventitious Shoot Regeneration of Oriental Lily (Lilium orientalis) and Genetic Stability Evaluation Based on ISSR Marker Variation. In Vitro Cell Development-Plant, 48(2), 172–179. https://doi.org/10.1007/s11627-012-9429-0
• Yadav, R., Yadav, N., Pal, M., Goutam, U. (2013). Multiple Shoot Proliferation, Bulblet İnduction and Evaluation of Genetic Stability in Asiatic Hybrid Lily (Lilium sp.). Indian Journal of Plant Physiology,18(4), 354–359. https://doi.org/10.1007/s40502-014-0060-4
• Yin, Z.F., Zhao, B., Bi, W.L., Chen, L., Wang, Q.C. (2013). Direct Shoot Regeneration From Basal Leaf Segments of Lilium and Assessment Of Genetic Stability in Regenerants by ISSR and AFLP Markers. In Vitro Cell Development-Plant, 49(3), 333–342. https://doi.org/10.1007/s11627-013-9501-4
• Bürün, B., Şahin, O. (2013). Micropropagation of Lilium candidum L.: A Rare and Native Bulbous Flower of Turkey. Bangladesh Journal of Botany, 42(2), 185–187. https://doi.org/10.3329/bjb.v42i1.15913
• Bi, W.L., Yin, Z.F., Guo, L., Chen, L., Pan, C., Wang, Q.C. (2015). Plant Regeneration From Shoot Regrowth and De Novo Embryo-Like Structures from Cryopreserved Shoot Tips of Lilium spp. In Vitro Cellular & Developmental Biology-Plant, 51(4), 390–398. https://doi.org/10.1007/s11627-015-9696-7
• Asmita, Sindhu, S.S., Jayanthi, M., Dhiman, M.R., Singh, M.K., Hossain, F. (2018). Genetic Fidelity Study of The in vitro Regenerated Plants in L.A. Hybrids of Lilium cv. Pavia. International Journal of Current Microbiology and Applied Sciences, 7(9), 1223–1231. https://doi.org/10.20546/ijcmas.2018.709.146
• Murashige, T., Skoog, F. (1962). A Revised Medium for Rapid Growth and Bio Assays with Tobacco Tissue Cultures. Physiologia Plantarum, 15(3), 473–497. https://doi.org/10.1111/j.1399-3054.1962.tb08052.x
• Rugini, E. (1993). In vitro Propagation of Some Olive Cultivars, Scientia Horticulturae, 24, 123–134. https://doi.org/10.1016/0304-4238(84)90143-2
• Kaya, E., Souza, F., Gökdoğan, E. Y., Ceylan, M., & Jenderek, M. (2017). Cryopreservation of Citrus Seed via Dehydration Followed by Immersion in Liquid Nitrogen. Turkish Journal of Biology, 41(1), 242-248.
• Marascuilo, L.A., McSweeney, M. (1977). Post-hoc Multiple Comparisons in Sample Preparations for Test of Homogenesity. In: McSweeney M, Marascuilo LA (eds) Non-parametric and Distribution Free Methods The Social Sciences. Books/Cole Publication, Belmont, pp 141–147.
• Lodhi, M.A., Ye, G.Y., Weeden, N.F., Reisch, B.I. (1994). A Simple and Efficient Method for DNA Extraction from Grapevine Cultivars and Vitis Species. Plant Molecular Biology Reporter, 12(1), 6–13. https://doi.org/10.1007/BF02668658
• Du, F., Wu, Y., Zhang, L., Li, X.W., Zhao, X.Y., Wang, W.H., Gao, Z.S., Xia, Y. (2015). De Novo Assembled Transcriptome Analysis and SSR Marker Development of a Mixture of Six Tissues from Lilium Oriental Hybrid ‘Sorbonne’. Plant Molecular Biology Reporter, 33, 281–293. https://doi.org/10.1007/s11105-014-0746-9
• George, E.F. (1993). Plant Propagation by Tissue Culture I: The Technology. Exegetics Ltd, Edington, UK.
• Loyola-Vargas, V.M., Ochoa-Alejo, N. (2012). An Introduction to Plant Cell Culture: The Future Ahead. Methods In Molecular Biology, Plant Cell Culture Protocols, Springer Protocols.
• Jeong, J.H. (1996). In Vitro Propagation of Bulb Scale Section of Several Korean Native Lilies. Acta Horticulturae (414), 269-276. https://doi.org/10.17660/ActaHortic.1996.414.33
• Varshney, A., Dhawan, V., Srivastava, P.S. (2000). A protocol for In vitro Mass Propagation of Asiatic Hybrids of Lily Through Liquid Stationary Culture. In vitro Cellular and Development Biology-Plant, 36(5), 383-391. https://doi.org/10.1007/s11627-000-0069-4
• Kumar, S., Sharma, D.R., Sharma, Y.D., Pathani, N.S. (2007). Influence of Growth Regulators and Nitrogenous Compounds on In Vitro Bulblet Formation And Growth in Oriental Lily. Horticultural Science, 34(2), 77-83.
• Saifullah, K., Sheeba, N., Mariam, R., Naheed, K., Asma, N., Bushra, S. (2010). Cultivation of Lilies (Lilium regale) for Commercialization in Pakistan. Pakistan Journal of Botany, 42(2), 1103-1113.
• Leshem, B., Lilien-Kipnis, H., Stenitz, B. (1982). The effect of light and of explant Oriention on The Regeneration and Subsequent Growth of Bulblets on Lilium longiflorum Thunb. Bulb-scale Sections Cultured in vitro. Scientia Horticulturae, 17, 129-136. https://doi.org/10.1016/0304-4238(82)90005-X
• Niimi, Y., Nakano, M., Isogai, N. (1999). Effects of Temperature and Illuminating Conditions on Regeneration and Development of Bulblets In Scale Culture Of Seven Lilium spp. Journal of The Japanese Society for Horticultural Science, 68(1), 28-34. https://doi.org/10.2503/jjshs.68.28
• Lian, M.L., Murthy, H.N., Paek, K.Y. (2002). Effects of Light Emitting Diodes (LEDs) on The In vitro Induction and Growth of Bulblets of Lilium oriental Hybrid ‘Pesaro’. Scientia Horticulturae, 94(3), 365-370. https://doi.org/10.1016/S0304-4238(01)00385-5
• Nhut, D.T., Huong, N.T.D., Le, B., Van, D.S., Jaime, T., Fukai, S., Tanaka, M. (2002). The Changes in Shoot Regeneration Potential of Protocorm-Like Bodies Derived from Lilium Iongiflorum Young Stem Explants Exposed to Medium Volume, pH, Light Intensity and Sucrose Concentration Pretreatment. The Journal of Horticultural Science and Biotechnology, 77(1), 79-82. https://doi.org/10.1080/14620316.2002.11511461
• Kumar, S., Kashyap, M., Sharma, D.R. (2005). In vitro Regeneration and Bulblet Growth From Lily Bulb Scale Explants as Affected by Retardants, Sucrose and Irradiance. Biologia Plantarum, 49(4), 629-632. https://doi.org/10.1007/s10535-005-0064-z
• Stimart, D. P., DP, S., PD, A. (1978). Tissue Culture of Bulb Scale Sections for Asexual Propagation of Lilium longiflorum Thunb. Journal of American Horticultural Science, 103(2), 182-184.
• Altan, F., Bürün, B. (2017). The Effect of Some Antibiotic And Fungucide Applications on the Micropropagation of Lilium candidum L. Mugla Journal of Science and Technology, 3(1), 86-91. https://doi.org/10.22531/muglajsci.307105
• Vinterhalter, B., Ninković, S., Zdravković, K.S., Subotić, A., Vinterhalter, D. (2007). Effect of Nitrogen Salts on The Growth of Ceratonia siliqua Shoot Cultures. Archives of Biological Sciences, 59(3), 217–222.
• McCoy, T.J., Smith, L.Y. (1986). Interspecific Hybridization Of Perennial Medicago Species Using Ovule-Embryo Culture. Theoretical and Applied Genetics, 71, 772–783. https://doi.org/10.1007/BF00276417
• Leljak-Levanic, D., Bauer, N., Mihaljevic´, S., Jelaska, S. (2004). Somatic Embryogenesis in Pumpkin (Cucurbita Pepo): Control of Somatic Embryo Development by Nitrogen Compounds. Journal of Plant Physiology, 161, 229–236. https://doi.org/10.1078/0176-1617-01055
• Altan, F., Bürün, B., Şahin, N. (2010). Fungal Contaminants Observed During Micropropagation of Lilium Candidum L. and The Effect of Chemotherapeutic Substances Applied After Sterilization. African Journal of Biotechnology, 9(7), 991–995.
• Saker, M., Bekheet, S., Taha, H., Fahmy, A., and Moursy, H. (2000). Detection of Somaclonal Variations in Tissue Culture Derived Date Palm Plants Using Isoenzyme Analysis and RAPD Fingerprints. Biologia Plantarum, 43 (3), 347–351. https://doi.org/10.1023/A:1026755913034
• Osena, G., Amugune, O.N., Nyaboga, E.N. (2017). Genetic Stability of Cassava Plants Regenerated Through Organogenesis Using Microsatellite Markers. Journal of Plant Sciences, 5(1), 19–28. https://doi.org/10.11648/j.jps.20170501.13