Advertisement

Biologia Plantarum

, Volume 52, Issue 2, pp 355–360 | Cite as

Direct organogenesis from leaf explants of Stevia rebaudiana and cultivation in bioreactor

  • R. V. Sreedhar
  • L. Venkatachalam
  • R. Thimmaraju
  • N. Bhagyalakshmi
  • M. S. Narayan
  • G. A. Ravishankar
Brief Communication

Abstract

Shoot buds were induced directly on either side of midrib from adaxial surface of immature leaf explants in Stevia rebaudiana Bertoni five weeks after culturing in Murashige and Skoog’s nutrient medium supplemented with 8.88 µM of N 6-benzylaminopurine and kinetin ranging from 4.65 to 6.98 µM. Immature leaves of 0.6 to 1 cm were found to produce best response (93 %) with a highest number of 4.93 shoot buds per explant. For elongation of regenerated shoot buds, MS medium supplemented with 30 g dm−3 sucrose and indole-3-butyric acid (IBA) ranging from 4.92 to 7.38 µM were found most suitable. The medium was further modified to suit bioreactor cultivation of regenerated shoots wherein the use of two-fold MS salts and 60 g dm−3 sucrose resulted in a high biomass yield of 50.68 g dm−3 (m/v) accounting for about 590 micro-cuttings in three weeks. Best rooting of micro-cuttings occurred in half strength MS medium supplemented with IBA ranging from 4.92 to 7.38 µM, 15 g dm−3 sucrose and gelled with 0.8 % agar. Rooted plants were successfully established in substrate containing sand, Vermicompost and garden soil in equal proportions and grown in greenhouse. This is the first report on direct shoot regeneration from Stevia leaves.

Additional key words

auxin cytokinin de novo shoot regeneration micro-cuttings micropropagation rooting 

Abbreviations

BA

N 6-benzylaminopurine

2,4-D

2,4-dichlorophenoxyacetic acid

IAA

indole-3-acetic acid

IBA

indole-3-butyric acid

Kn

kinetin

M

Murashige and Skoog

Preview

Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.

References

  1. Akita, M., Shigeoka, T., Koizumi, Y., Kawamura, M.: Mass propagation of shoots of Stevia rebaudiana using a large scale bioreactor.-Plant Cell Rep. 13: 180–183, 1994.Google Scholar
  2. Bespalhok, J.C., Vieira, L.G.E., Hashimoto, J.M.: Factores influenciando a micropropagacao in vitro de gemas axilares de Stevia rebaudiana.-Bertoni R Bras. Fisiol. Veg. 4: 59–61, 1992.Google Scholar
  3. Famiani, F., Ferradini, N., Staffolani, P., Standardi, A.: Effect of leaf excision time and age, BA concentration and dark treatments on in vitro shoot regeneration of M 26 apple rootstocks.-J. hort. Sci. 69: 679–685, 1994.Google Scholar
  4. Ferreira, C.M., Handro, W.: Production, maintenance and plant regeneration from cell suspension cultures of Stevia rebaudiana Bertoni.-Plant Cell Rep. 7: 123–126, 1988.CrossRefGoogle Scholar
  5. Flachsland, E., Mroginski, L., Davina, J.: Regeneration of plants from anther of Stevia rebaudiana Bertoni (Compositae) cultivated in vitro.-BioCell 20: 87–90, 1996.Google Scholar
  6. Harter, L.N.: Critical values for Duncan are new multiple range test.-Biometrics 16: 671–685, 1960.CrossRefGoogle Scholar
  7. Ibrahim, R., Pierre, C.D.: Factors controlling high efficiency adventitious bud formation and plant regeneration from in vitro leaf explants of roses (Rosa hybrida L.).-Sci. Hort. 88: 41–57, 2001.CrossRefGoogle Scholar
  8. Karam, N.S., Al-Majathoup, M.: Direct shoot regeneration and microtuberization in wild Cyclamen persicum Mill. using seedling tissue.-Sci. Hort. 86: 235–236, 2000.CrossRefGoogle Scholar
  9. Kumar, P.P., Dimps Rao, C., Goh, C.J.: Influence of petiole and lamina on adventitious shoot initiation from leaf explants of Paulownia fortunei.-Plant Cell Rep. 17: 886–890, 1998.CrossRefGoogle Scholar
  10. Murashige, T., Skoog, F.A.: Revised medium for rapid growth and bioassays with tobacco tissue cultures.-Physiol. Plant. 15:473–497, 1962.CrossRefGoogle Scholar
  11. Sivaram, L., Mukundan, U.: In vitro culture studies on Stevia rebaudiana.-In vitro cell. dev. Biol. Plant. 39: 520–523, 2003.CrossRefGoogle Scholar
  12. Takayama, S., Akita, M.: The types of bioreactors used for shoots and embryos.-Plant Cell Tissue Organ Cult. 39:147–156, 1994.CrossRefGoogle Scholar
  13. Tamura, T., Nakamura, S., Fukui, H., Tabata, M.: Clonal propagation of Stevia rebaudiana Bertoni by stem-tip culture.-Plant Cell Rep. 3: 183–185, 1984.CrossRefGoogle Scholar
  14. Thimmaraju, R., Venkatachalam, L., Vinod Kumar, Bhagyalakshmi, N., Sreedhar, R.V., Ravishankar, G.A.: Peroxidase production from hairy root cultures of red beet (Beta vulgaris).-J. Biotechnol. 8: 185–196, 2005.Google Scholar
  15. Wang, H.-M., Zu Y.-G., Wang, W.-J., Dong, F.-L.: Establishment of Camptotheca acuminata regeneration from leaf explants.-Biol. Plant. 50: 725–728, 2006.CrossRefGoogle Scholar
  16. Welander, M., Maheswaran, G.: Shoot regeneration from leaf explants of dwarfing apple rootstocks.-J. Plant Physiol. 140: 223–228, 1992.Google Scholar

Copyright information

© Institute of Experimental Botany, Academy of Sciences of the Czech Republic, Praha 2008

Authors and Affiliations

  • R. V. Sreedhar
    • 1
  • L. Venkatachalam
    • 1
  • R. Thimmaraju
    • 1
  • N. Bhagyalakshmi
    • 1
  • M. S. Narayan
    • 1
  • G. A. Ravishankar
    • 1
  1. 1.Plant Cell Biotechnology DepartmentCentral Food Technological Research InstituteMysoreIndia

Personalised recommendations