Advertisement

Sugar Tech

pp 1–9 | Cite as

Agrobacterium rhizogenes-Mediated Transformation Enhances Steviol Glycosides Production and Growth in Stevia rebaudiana Plantlets

  • Ángel de Jesús Sanchéz-Cordova
  • Jacqueline Capataz-Tafur
  • Blanca Estela Barrera-Figueroa
  • Adolfo López-Torres
  • Paul Mauricio Sanchez-Ocampo
  • Edgar García-LópezEmail author
  • Ariana Arlene Huerta-HerediaEmail author
Research Article
  • 21 Downloads

Abstract

Stevia rebaudiana accumulates steviol glycosides (SGs) that are used as noncaloric, natural sweeteners in food industry. The most important SGs are rebaudioside A (Reb A) and stevioside (St), and they are markers of the quality of S. rebaudiana lines. In this work, the production and physiology of SGs in Agrobacterium rhizogenes-transformed plantlets were analyzed. By means of HPLC–DAD, the production of St and Reb A was quantified, resulting in a 1.4- and 1.5-fold production increase of St and Reb A, respectively, in transformed compared to wild-type plantlets. Superior phenotype of transformed plantlets was observed in comparison with wild type, represented as an increase of 25% in biomass of aerial parts, 43% in biomass of roots, 20–30% of leaf area and 24% of chlorophylls content. Moreover, the SG production profiles evaluated in a 20-d period showed higher yield in the transformed line. The results of this work demonstrated the usefulness of S. rebaudiana transformation via A. rhizogenes as a strategy to explore new approaches for the improvement of SGs production.

Keywords

Stevia rebaudiana Stevioside Genetic transformation Rebaudioside A Agrobacterium 

Notes

Acknowledgements

The authors are grateful to the National Council of Science and Technology (Consejo Nacional de Ciencia y Tecnología—CONACyT) for scholarship No. 615648 to AJSC and to Catedras-CONACyT 3212 (235307) and INFR201501 (255514).

Author’s Contributions

A.J.S-C carried out laboratory work and data analysis. J.C-T contributed to the equipment and B.E.B-F edited the manuscript. P.M.S-O. and A.L-T. contributed to the data interpretation and statistical analysis. A.A.H-H. and E.G-L. designed and analyzed the experiments and wrote the manuscript. All authors reviewed and approved the final manuscript.

Compliance with Ethical Standards

Conflict of interest

The authors declare that they have no conflict of interest.

References

  1. Amselem, J., and M. Tepfer. 1992. Molecular basis of novel root phenotypes induced by Agrobacterium rhizogenes A4 on cucumber. Plant Molecular Biology 19: 421–432.CrossRefGoogle Scholar
  2. Anbazhagan, M., M. Kalpana, R. Rajendran, V. Natarajan, and D. Dhanavel. 2010. In vitro production of Stevia rebaudiana Bertoni. Emirates Journal of Food Agriculture 2: 216–222.CrossRefGoogle Scholar
  3. Bayraktar, M., E. Naziri, I. Hakki, A. Fatih, K. Esra, I. Begum, A. Erdal, and B.A. Gurel. 2016. Elicitor induced stevioside production, in vitro shoot growth, and biomass accumulation in micropropagated Stevia rebaudiana. Plant Cell, Tissue and Organ Culture 127: 289–300.CrossRefGoogle Scholar
  4. Bender, C., S. Graziano, and B.F. Zimmermann. 2015. Study of Stevia rebaudiana Bertoni antioxidant activities and cellular properties. International Journal of Food Sciences and Nutrition 66: 553–558.CrossRefGoogle Scholar
  5. Bonhomme, V., D. Laurain-Mattar, and M.A. Fliniaux. 2000. Effects of the rolC gene on hairy root: Induction development and tropane alkaloid production by Atropa belladonna. Journal of Natural Products 63: 1249–1252.CrossRefGoogle Scholar
  6. Bulgakov, V.P. 2008. Functions of rol genes in plant secondary metabolism. Biotechnolgy Advances 26: 318–324.CrossRefGoogle Scholar
  7. Calderón-Gabriel, L., A. Jiménez-Brigada, A.A. Huerta-Heredia, J. Capataz-Tafur, and E. García-López. 2016. Effect of three strains of Agrobacterium rhizogenes and explant type on genetic transformation of Stevia rebaudiana. Mexican Journal of Biotechnology 1: 34–41.Google Scholar
  8. Choi, P.S., Y.D. Kim, K.M. Choi, H.J. Chung, D.W. Choi, and J.R. Liu. 2004. Plant regeneration from hairy-root cultures transformed by infection with Agrobacterium rhizogenes in Catharanthus roseus. Plant Cell Reports 22: 828–831.CrossRefGoogle Scholar
  9. Christey, M.C. 2001. Use of Ri-mediated transformation for production of transgenic plants. Vitro Cellular & Developmental Biology-Plant 37: 687–700.CrossRefGoogle Scholar
  10. David, C., M.D. Chilton, and J. Tempé. 1984. Conservation of T-DNA in plants regenerated from hairy root cultures. Nature Biotechnology 2: 73–76.CrossRefGoogle Scholar
  11. Dilshad, E., H. Ismail, R.M. Cusido, J. Palazon, K. Ramirez-Estrada, and B. Mirza. 2016. Rol genes enhance the biosynthesis of antioxidants in Artemisia carvifolia Buch. BMC Plant Biology 16: 125.CrossRefGoogle Scholar
  12. Edwards, K., C. Johnstone, and C. Thompson. 1991. A simple and rapid method for the preparation of plant genomic DNA for PCR analysis. Nucleic Acids Research 19: 1349.CrossRefGoogle Scholar
  13. Fallah, F., F. Nokhasi, M. Ghaheri, D. Kahrizi, A. Beheshti Ale Agha, T. Ghorbani, E. Kazemi, and Z. Ansarypour. 2017. Effect of salinity on gene expression, morphological and biochemical characteristics of Stevia rebaudiana Bertoni under in vitro conditions. Cellular and Molecular Biology 63: 102–106.CrossRefGoogle Scholar
  14. Fu, X., Z.P. Yin, J.G. Chen, X.C. Shangguan, X. Wang, Q.F. Zhang, and D.Y. Peng. 2015. Production of chlorogenic acid and its derivatives in hairy root cultures of Stevia rebaudiana. Journal Agricultural and Food Chemistry 63: 262–268.CrossRefGoogle Scholar
  15. Gamboa, F., and M. Chaves. 2012. Antimicrobial potential of extracts from Stevia rebaudiana leaves against bacteria of importance in dental caries. Acta Odontológica Latinoamericana 25: 171–175.PubMedGoogle Scholar
  16. Ghorbani, T., D. Kahrizi, M. Saeidi, and I. Arji. 2017. Effect of sucrose concentrations on Stevia rebaudiana Bertoni tissue culture and gene expression. Cellular and Molecular Biology 63: 33–37.CrossRefGoogle Scholar
  17. Gunjan, S.K., J. Lutz, A. Bushong, D.T. Rogers, and J. Littleton. 2013. Hairy root cultures and plant regeneration in Solidago nemoralis transformed with Agrobacterium rhizogenes. American Journal of Plant Sciences 4: 1675–1678.CrossRefGoogle Scholar
  18. Gupta, E., S. Kaushik, S. Purwar, R. Sharma, A.K. Balapure, and S. Sundaram. 2017. Anticancer potential of steviol in MCF-7 human breast cancer cells. Pharmacognosy Magazine 13: 345–350.CrossRefGoogle Scholar
  19. Gupta, E., S. Purwar, S. Sundaram, and G.K. Rai. 2013. Nutritional and therapeutic values of Stevia rebaudiana: A review. Journal of Medicinal Plants Research 7: 3343–3353.Google Scholar
  20. Gurusamy, P.D., H. Schäfer, S. Ramamoorthy, and M. Wink. 2017. Biologically active recombinant human erythropoietin expressed in hairy root cultures and regenerated plantlets of Nicotiana tabacum L. PLoS ONE 12 (8): e0182367.CrossRefGoogle Scholar
  21. Ishizaki, T., Y. Hoshino, K. Masuda, and K. Oosawa. 2002. Explants of Ri-transformed hairy roots of spinach can develop embryogenic calli in the absence of gibberellic acid, an essential growth regulator for induction of embryogenesis from non-transformed roots. Plant Science 163: 223–231.CrossRefGoogle Scholar
  22. Javed, R., A. Mohamed, B. Yücesan, E. Gurel, R. Kausar, and M. Zia. 2017a. CuO nanoparticles significantly influence in vitro culture, steviol glycosides, and antioxidant activities of Stevia rebaudiana Bertoni. Plant Cell, Tissue and Organ Culture 131: 611–620.CrossRefGoogle Scholar
  23. Javed, R., M. Usman, B. Yücesan, M. Zia, and E. Gürel. 2017b. Effect of zinc oxide (ZnO) nanoparticles on physiology and steviol glycosides production in micropropagated shoots of Stevia rebaudiana Bertoni. Plant Physiology and Biochemistry 110: 94–99.CrossRefGoogle Scholar
  24. Khan, S.A., L.U. Rahman, K. Shanker, and M. Singh. 2014. Agrobacterium tumefaciens mediated transgenic plant and somaclone production through direct and indirect regeneration from leaves in Stevia rebaudiana with their glycoside profile. Protoplasma 251: 661–670.CrossRefGoogle Scholar
  25. Khan, S.A., L.U. Rahman, R. Verma, and K. Shanker. 2016. Physical and chemical mutagenesis in Stevia rebaudiana: Variant generation with higher UGT expression and glycosidic profile but with low photosynthetic capabilities. Acta Physiologiae Plantarum 38: 1–12.CrossRefGoogle Scholar
  26. Ladygin, V.G., N.I. Bondarev, G.A. Semenova, A.A. Smolov, O.V. Reshetnyak, and A.M. Nosov. 2008. Chloroplast ultrastructure, photosynthetic apparatus activities and production of steviol glycosides in Stevia rebaudiana in vivo and in vitro. Biologia Plantarum 52: 9–16.CrossRefGoogle Scholar
  27. Libik-Konieczny, M., E. Capecka, E. Kąkol, M. Dziurka, A. Grabowska-Joachimiak, E. Sliwinska, and L. Pistelli. 2018. Growth, development and steviol glycosides content in the relation to the photosynthetic activity of several Stevia rebaudiana Bertoni strains cultivated under temperate climate conditions. Scientia Horticulturae 234: 10–18.CrossRefGoogle Scholar
  28. Lichtenthaler, H.K., and A.R. Wellburn. 1983. Determinations of total carotenoids and chlorophylls a and b of leaf extracts in different solvents. Biochemical Society Transactions 11: 591–592.CrossRefGoogle Scholar
  29. López-Laredo, A.R., F.D. Ramírez-Flores, G. Sepúlveda-Jiménez, and G. Trejo-Tapia. 2009. Comparison of metabolite levels in callus of Tecoma stans (L.) Juss. ex Kunth. cultured in photoperiod and darkness. Vitro Cellular & Developmental Biology-Plant 45: 550–558.CrossRefGoogle Scholar
  30. Magangana, T.P., M.A. Stander, and N.P. Makunga. 2018. Effect of nitrogen and phosphate on in vitro growth and metabolite profiles of Stevia rebaudiana Bertoni (Asteraceae). Plant Cell, Tissue and Organ Culture 134: 141–151.CrossRefGoogle Scholar
  31. Matveeva, T.V., S.V. Sokornova, and L.A. Lutova. 2015. Influence of Agrobacterium oncogenes on secondary metabolism of plants. Phytochemistry Reviews 14: 541–554.CrossRefGoogle Scholar
  32. Murashige, T., and F. Skoog. 1962. A revised medium for rapid growth and bioassays with tobacco tissue cultures. Physiologia Plantarum 15: 473–497.CrossRefGoogle Scholar
  33. Namdari, N., L. Shooshtari, and A. Qaderi. 2015. In vitro micropropagation of Stevia rebaudiana Bertoni. Biological Forum—An International Journal 7: 1750–1754.Google Scholar
  34. Pandey, H., P. Pandey, S.S. Pandey, S. Singh, and S. Banerjee. 2016. Meeting the challenge of stevioside production in the hairy roots of Stevia rebaudiana by probing the underlying process. Plant Cell, Tissue and Organ Culture 126: 511–521.CrossRefGoogle Scholar
  35. Ramos-Tovar, E., E. Hernández-Aquino, S. Casas-Grajales, L.D. Buendia-Montaño, S. Galindo-Gómez, J. Camacho, V. Tsutsumi, and P. Muriel. 2018. Stevia prevents acute and chronic liver injury induced by carbon tetrachloride by blocking oxidative stress through Nrf2 upregulation. Oxidative Medicine Cellular Longevity 2018: 3823426.  https://doi.org/10.1155/2018/3823426.CrossRefPubMedGoogle Scholar
  36. Razak, U.N.A.A., C.B. Ong, T.S. Yu, and L.K. Lau. 2014. In vitro micropropagation of Stevia rebaudiana Bertoni in Malaysia. Brazilian Archives of Biology Technology 57: 23–28.CrossRefGoogle Scholar
  37. Roychowdhury, D., A. Majumder, and S. Jha. 2013. Agrobacterium rhizogenes-mediated transformation in medicinal plants: Prospects and challenges. In Biotechnology for medicinal plants: Micropropagation and improvement, ed. S. Chandra, H. Lata, and A. Varma, 29–68. Berlin/Heidelberg: Springer.CrossRefGoogle Scholar
  38. Ruiz-Ruiz, J.C., Y.B. Moguel-Ordoñez, and M.R. Segura-Campos. 2017. Biological activity of Stevia rebaudiana Bertoni and their relationship to health. Critical Reviews in Food Science and Nutrition 57: 2680–2690.CrossRefGoogle Scholar
  39. Sarkar, S., I. Ghosh, D. Roychowdhury, and S. Jha. 2018. The Effects of rol genes of Agrobacterium rhizogenes on morphogenesis and secondary metabolite accumulation in medicinal plants. In Biotechnological approaches for medicinal and aromatic plants, ed. N. Kumar, 27–51. Singapore: Springer.CrossRefGoogle Scholar
  40. Shkryl, Y.N., G.N. Veremeichik, V.P. Bulgakov, G.K. Tchernoded, N.P. Mischenko, S.A. Fedoreyev, and Y.N. Zhuravlev. 2007. Individual and combined effects of the rolA, B and C genes on anthraquinone production in Rubia cordifolia transformed calli. Biotechnology and Bioengineering 100: 118–125.CrossRefGoogle Scholar
  41. Tada, A., K. Ishizuki, J. Iwamura, H. Mikami, Y. Hirao, I. Fujita, T. Yamazaki, H. Akiyama, and Y. Kawamura. 2013. Improvement of the assay method for steviol glycosides in the JECFA specifications. American Journal of Analytical Chemistry 4: 190–196.CrossRefGoogle Scholar
  42. Tepfer, D. 1984. Genetic transformation of several species of higher plants by Agrobacterium rhizogenes: Phenotypic consequences and sexual transmission of transformed genotype and phenotype. Cell 37: 959–967.CrossRefGoogle Scholar
  43. Tzfira, T., and V. Citovsky. 2006. Agrobacterium-mediated genetic transformation of plants: Biology and biotechnology. Current Opinion in Biotechnology 17: 147–154.CrossRefGoogle Scholar
  44. Yadav, S.K., and P. Guleria. 2012. Steviol glycosides from Stevia: Biosynthesis pathway review and their application in foods and medicine. Critical Reviews in Food Science and Nutrition 52: 988–998.CrossRefGoogle Scholar
  45. Yamazaki, T., H.E. Flores, K. Shimomura, and K. Yoshihira. 1991. Examination of steviol glucosides production by hairy root and shoot cultures of Stevia rebaudiana. Journal of Natural Products 54: 986–992.CrossRefGoogle Scholar
  46. Yang, D.C., and Y.E. Choi. 2000. Production of transgenic plants via Agrobacterium rhizogenes mediated transformation of Panax ginseng. Plant Cell Reports 19: 491–496.CrossRefGoogle Scholar
  47. Zhou, M.L., X.M. Zhu, J.R. Shao, Y.X. Tang, and Y.M. Wu. 2011. Production and metabolic engineering of bioactive substances in plant hairy root culture. Applied Microbiology and Biotechnology 90: 1229–1239.CrossRefGoogle Scholar

Copyright information

© Society for Sugar Research and Promotion 2019

Authors and Affiliations

  1. 1.División de Estudios de PosgradoUniversidad del PapaloapanTuxtepecMexico
  2. 2.Instituto de BiotecnologíaUniversidad del PapaloapanTuxtepecMexico
  3. 3.Instituto de Química AplicadaUniversidad del PapaloapanTuxtepecMexico
  4. 4.Cátedras CONACyT-UNPAUniversidad del PapaloapanTuxtepecMexico

Personalised recommendations