Optimization of factors affecting in vitro regeneration, flowering, ex vitro rooting and foliar micromorphological studies of Oldenlandia corymbosa L.: a multipotent herb
- 27 Downloads
The conditions were optimized for efficient in vitro regeneration of shoots and roots of Oldenlandia corymbosa L. using nodal shoot explants. Murashige and Skoog’s (MS) medium augmented with additives and 2.0 mg L−1 BAP was recorded optimum for shoot bud induction from the nodal meristems. The shoots were proliferated by subsequent subcultures on half strength MS medium fortified with 1.0 mg L−1 BAP and 0.5 mg L−1 kinetin + additives. This media combination yielded maximum number of shoots (223 ± 4.12 shoots/culture bottle) with 13.4 cm average length. Flower buds were induced (4.2 ± 0.28 flowers) from the in vitro multiplied shoots on MS medium contained 1.0 mg L−1 BAP and 0.5 mg L−1 of kinetin and IAA under 50 µmol m−2 s−1 SFPD light intensity for 12 h/day photoperiod. In vitro regenerated shoots were rooted on half strength MS medium conjunct with IAA, IBA and NAA singly at different concentrations. The best rooting response was observed on half strength MS medium containing IBA at 2.0 mg L−1 with activated charcoal. Roots were also induced from the cut ends of the shoots using ex vitro rooting techniques in O. corymbosa by pulse treating the shoots with 300 mg L−1 IBA for 4 min. Better roots were achieved in this method than the in vitro roots in terms of numbers (14.7 ± 0.21) and firmness. The foliar micromorphological studies could help to understand the structural adaptations of micropropagated O. corymbosa plantlets towards field environments. The in vitro induced anomalies in stomatal apparatus were repaired during hardening of plantlets in the greenhouse and after field transfer. Decrease in stomatal density (from 70.78 ± 0.55 to 55.6 ± 0.10), and increase in veins, trichomes and crystals/raphides densities revealed the developments of structural changes in the leaves to withstand in the harsh field conditions. The acclimatized plantlets with well developed root systems were successfully shifted to the natural soils with 98% survival rate.
KeywordsIn vitro regeneration Oldenlandia corymbosa Flower induction Ex vitro rooting Acclimation Micromorphological studies
Murashige and Skoog’s basal medium
α-Naphthalene acetic acid
Spectral photon flux density
Authors are grateful to the Department of Science, Technology and Environment, Government of Puducherry, India for providing financial support to their laboratory under the Grant-In-Aid Scheme.
JR performed the experiments and analyzed data. MM designed the experiment and contributed to writing the manuscript. MSS conceived the idea, and contributed to writing and editing the manuscript.
Compliance with ethical standards
Conflict of interest
The authors declare that they have no conflict of interest.
The authors declare that the study was carried out following accepted professional conduct. However no ethical approval was needed for the study as it did not involve the use of animals or human subjects.
- Attims Y, Come D (1978) Dormance des graines d’une plante tropicale (Oldenlandia corymbosa L, Rubiacees): Selection de deux types de plantes. CR Acad Sci Ser D 286:1669–1672Google Scholar
- Chen W, Zou SQ, Li KQ (2005) Extracting and identifying of ursolic acid from Hedyotis corymbosa Lam. Jiangxi Shifan Daxue Xuebao, Ziran Kexueban 29:126–128Google Scholar
- Deng ZC, Jin H, He H (2015) An efficient micropropagation system for Morinda officinalis How. (Rubiaceae), an endangered medicinal plant. J Agric Sci Tech 17:1609–1618Google Scholar
- Do Cao T, Attims Y, Corbineau F, Come D (1978) Germination des graines produites par les plantes de deux lignees d’Oldenlandia corymbosa L. (Rubiacees) cultiviees dans des conditions controlees. Physiol Veg 16:521–531Google Scholar
- Dutta R, Deb DB (2004) Taxonomic revision of Hedyotis L. (Rubiaceae) in Indian subcontinent. Botanical Survey of India, Kolkata, pp 1–211Google Scholar
- Gamble JS (1921) Flora of the Presidency of Madras, vol 1, London. Repr edn. (1995) Bishen Singh, Mahendra pal singh, DehradunGoogle Scholar
- Ghani A (2003) Medicinal plants of Bangladesh, 2nd edn. Asiatic society of Bangladesh, DhakaGoogle Scholar
- Johansen DA (1940) Plant microtechnique, 1st edn. McGraw Hill Book Co, New York, pp 182–197Google Scholar
- Joshi A, Mathur N (2015) In vitro propagation and conservation of Anthocephalus cadamba through apical bud and nodal explants—a valuable medicinal plant. CIBTech J Biotechnol 4(3):8–18Google Scholar
- Kanechi M, Ochi M, Abe M, Inagaki N, Maekawa S (1998) The effects of carbon dioxide enrichment, natural ventilation, and light intensity on growth, photosynthesis, and transpiration of cauliflower plantlets cultured in vitro photoautotrophically and photomixotrophically. J Am Soc Hortic Sci 123:176–181Google Scholar
- Lai KD, Tran VS, Pham GD (2002) Two anthraquinones from Hedyotis corymbosa and Hedyotis diffusa. Tap Chi Hoa Hoc 40:66–68Google Scholar
- Patel AK, Lodha D, Ram K, Shekhawat S, Shekhawat NS (2016) Evaluation of physiochemical factors affecting high-frequency plant regeneration of Blyttia spiralis (synonym: Pentatropis spiralis), a threatened climber of medicinal value. In Vitro Cell Dev Biol-Plant. https://doi.org/10.1007/s11627-015-9738-1 Google Scholar
- Permadi A (2006) Urine facilitating medicinal Plants. Sower Self Reliance, LondonGoogle Scholar
- Prabhakar M (2004) Structure, delimitation, nomenclature and classification of stomata. Acta Bot Sin 46:242–252Google Scholar
- Salisbury EJ (1932) The interrelations of soil climate and organisms and the use of stomatal frequency as an integrating index of relation of the plant. Bech Bot Zbl 99:402–420Google Scholar
- Shekhawat MS, Kannan N, Manokari M, Revathi J (2012) In vitro propagation of Oldenlandia umbellata L.—a highly medicinal & dye-yielding plant of coromandel coast. Int J Recent Sci Res 3(9):758–761Google Scholar
- Tisserat B, Galletta PD (1995) In vitro flowering and fruiting of Capsicum frutescens L. Hortic Sci 30:130–132Google Scholar