Abstract
This study describes for the first time a simple and quick direct somatic embryogenesis and plant regeneration protocol for turmeric (Curcuma longa L.) using leaf base explants. Preincubation of explants was carried out on solid Murashige and Skoog (MS) medium containing 4.49 μM 2,4-dichlorophenoxy acetic acid for 20 days under continuous dark condition. Subsequently transfer of the explants for 2 weeks to MS liquid medium containing 1.32 μM 6-benzyladenine (BA) under illuminated condition resulted in the highest percentage of somatic embryo (91.1 %) induction. Secondary somatic embryos which originated from primary somatic embryos were also observed in the same medium supplemented with 2.20 and 2.64 μM BA. Thereafter, mature somatic embryos germinated readily and the maximal plantlet development (87 %) was achieved on 1/2-strength MS medium containing gibberellic acid (GA3) under dark condition. Histological analysis revealed that the proembryogenic masses were initiated directly from the area near the vascular tissues of the leaf base explants. Presence of clear protoderm in the globular structure and procambial strands in the matured stage of somatic embryos confirmed that these structures were true somatic embryos. Somatic embryos were successfully acclimatized ex vitro at a survival rate of 72.9 % and they were normal phenotypes.
Similar content being viewed by others
Abbreviations
- 2,4-D:
-
2,4-dichlorophenoxyacetic acid
- BA:
-
6-benzyladenine
- IBA:
-
Indole-3-butyric acid
- GA3 :
-
Gibberellic acid
- MS:
-
Murashige and Skoog medium (1962)
- SEM:
-
Scanning electron microscopy
References
Abohatem M, Zouine J, Hadrami IE (2011) Low concentrations of BAP and high rate of subcultures improve the establishment and multiplication of somatic embryos in date palm suspension cultures by limiting oxidative browning associated with high levels of total phenols and peroxidase activities. Sci Hortic 130:344–348
Baba AI, Nogueira FCS, Pinheira CB, Brasil JN, Jereissati ES, Jucá TL, Soares AA, Santos MF, Domont GB, Campos FAP (2008) Proteome analysis of secondary somatic embryogenesis in cassava (Manihot esculenta). Plant Sci 175:717–723
Ball ST, Zhou H, Konzak CF (1993) Influence of 2,4-D, IAA, and duration of callus induction in anther culture of spring wheat. Plant Sci 90:195–200
Borkind C, Choi JH, Jin Z, Franz G, Hatzopoupos P, Chorneaau R, Bonas U, Pelegri F, Sung ZR (1988) Developmental regulation of embryogenic genes in plants. Proc Natl Acad Sci USA 85:6399–6503
Buffard-Morel J, Verdeil JL, Pannetier C (1992) Embryogenese somatique du cocotier (Cocos nucifera L.) a partir d’explants foliaires: etudes histologiques. Can J Bot 70:735–741
Cantelmo L, Soares BO, Rocha LP, Pettinelli JA, Callado CH, Mansur E, Castellar A, Gagliardi RF (2013) Repetitive somatic embryogenesis from leaves of the medicinal plant Petiveria alliacea L. Plant Cell Tiss Organ Cult 115:385–393
Cheethaparambil A, Pillai GS, Balachandran I (2013) Comparative studies on in vitro microrhizomeinduction in three varieties of Curcuma longa (Turmeric)—The role of two stress hormones. Int J Sci Res Manag 1:230–237
Choi YE, Kim JW, Yoon ES (1999a) High-efficiency plant production via somatic embryogenesis from callus or cell suspension cultures in Eleutherococcus senticosus. Ann Bot 83:309–314
Choi YE, Yang DC, Yoon ES (1999b) Rapid propagation of Eleutherococcus senticosus via direct somatic embryogenesis from explants of seedlings. Plant Cell Tiss Organ Cult 58:93–97
Cousins M, Adelberg J, Chen F, Rieck J (2007) Antioxidant capacity of fresh and dried rhizomes from four clones of turmeric (Curcuma longa L.) grown in vitro. Ind Crop Prod 25:129–135
Dam A, Paul S, Bandyopadhyay TK (2010) Direct somatic embryogenesis and plant regeneration from leaf explants of Limonium sinensis (Girard) Kuntze. Sci Hortic 126:253–260
Delgado-Vargas F, Jiménez-Aparicio AR, Paredes-López O (2000) Natural pigments: carotenoids, anthocyanins and betalains—characteristics, biosynthesis, processing and stability. CRC Crit Rev Food Sci Nut 40:173–289
Deo PC, Harding RH, Taylor M, Tyagi AP, Becker DK (2009) Somatic embryogenesis, organogenesis and plant regeneration in taro (Colocasia esculenta var. esculenta). Plant Cell Tiss Organ Cult 99:61–71
Downham A, Collins P (2000) Coloring our foods in the last and next millennium. Int J Food Sci Technol 35:5–22
Etienne H, Montoro P, Michaux-Ferriere N, Carron MP (1993) Effects of desiccation, medium osmolarity and abscisic acid on the maturation of Hevea brasiliensis somatic embryos. J Exp Bot 44:1613–1619
Feher A, Pasternak TP, Dudits D (2003) Transition of somatic cells to an embryogenic state. Plant Cell Tiss Organ Cult 74:201–228
Franklin G, Arvinth S, Sheeba CJ, Kanchana M, Subramonian N (2006) Auxin pretreatment promotes regeneration of sugarcane (Saccharum spp. hybrids) midrib segment explants. Plant Growth Regul 50:111–119
George EF, Hall MA, De Klerk GJ (eds) (2008) Plant propagation by tissue culture. The background, vol 1, 3rd edn. Springer, Dordrecht
Guo Y, Zhang Z (2005) Establishment and plant regeneration of somatic embryogenic cell suspension cultures of the Zingiber officinale Rosc. Sci Hortic 107:90–96
Halperin W (1964) Morphogenetic studies with partially synchronized cultures of carrot embryos. Science 146:408–410
He R, Gang DR (2013) Somatic embryogenesis and Agrobacterium-mediated transformation of turmeric (Curcuma longa). Plant Cell Tiss Organ Cult. doi:10.1007/s11240-013-0407-y
Jalil M, Chee WW, Othman RY, Khalid N (2008) Morphohistological examination on somatic embryogenesis of Musa acuminata cv. Mas (AA). Sci Hortic 117:335–340
John CK, Nadgauda RS, Mascarenhas AF (1997) Turmeric. Tissue culture of economic plants. Center for Science and Technology of the Non-Aligned and Other Developing Countries, New Delhi, pp 191–209
Kar B, Kuanar A, Singh S, Mohanty S, Joshi RK, Subudhi E, Nayak S (2013) In vitro induction, screening and detection of high essential oil yielding somaclones in turmeric (Curcuma longa L.). Plant Growth Regul. doi:10.1007/s10725-013-9836-1
Kuanar A, Kar B, Acharya L, Nayak S (2012) Nuclear DNA, DNA finger printing and essential oil content variation in callus derived regenerants of Curcuma longa L. Nucleus 55:101–106
Liang GH, Xu A, Tang H (1987) Direct generation of wheat haploids via anther culture. Crop Sci 27:336–339
Marsolais AA, Kasha KJ (1985) Callus induction from barley microspores: the role of sucrose and auxin in a barley anther culture medium. Can J Bot 63:2209–2212
Murashige T, Skoog F (1962) A revised medium for rapid growth and bioassay with tobacco tissue culture. Physiol Plant 15:473–497
Nayak S, Kaur T, Mohanty S, Ghosh G, Choudhury R, Acharya L, Subudhi E (2011) In-vitro and ex-vitro evaluation of long term micropropagated turmeric as analyzed through cytophotometry, phytoconstituents, biochemical and molecular markers. Plant Grow Regul 64:91–98
Nhut DT, Le BV, Van KTT (2000) Somatic embryogenesis and direct shoot regeneration of rice (Oryza sativa L.) using thin cell layer culture of apical meristematic tissue. J Plant Physiol 157:559–565
Panaia M, Bunn E, McComb J (2011) Primary and repetitive secondary somatic embryogenesis of Lepidosperma drummondii (Cyperaceae) and Baloskion tetraphyllum (Restionaceae) for land restoration and horticulture. In Vitro Cell Dev Biol Plant 47:379–386
Quiroz-Figueroa FR, Fuentes-Cerda CFJ, Rojas-Herrera R, Loyola-Vargas VM (2002) Histological studies on the developmental stages and differentiation of two different somatic embryogenesis systems of Coffea arabica. Plant Cell Rep 20:1141–1149
Salvi ND, George L, Eapen S (2000) Direct regeneration of shoots from immature inflorescence cultures of turmeric. Plant Cell Tiss Organ Cult 62:235–238
Salvi ND, George L, Eapen S (2001) Plant regeneration from leaf base callus of turmeric and random amplified polymorphic DNA analysis of regenerated plants. Plant Cell Tiss Organ Cult 66:113–119
Salvi ND, George L, Eapen S (2002) Micropropagation and field evaluation of micropropagated plants of turmeric. Plant Cell Tiss Organ Cult 68:143–151
Sané D, Aberlenc-Bertossi F, Gassama-Dia YK, Sagna M, Trouslot MF, Duval Y, Borgel A (2006) Histological analysis of callogenesis and somatic embryogenesis from cell suspensions of date palm (Phoenix dactylifera). Ann Bot 98:301–308
Satyavati GV, Raina MK, Sharma M (eds) (1976) Medicinal plants of India, vol I. Indian Council of Medical Research, New Delhi, pp 312–317
Schwandiman J, Pannetier C, Michaux-Ferriere N (1988) Histology of somatic embryogenesis from leaf explants of the oil palm Elaeis guineensis. Ann Bot 62:43–52
Sharma SK, Millam S (2004) Somatic embryogenesis in Solanum tuberosum L.: a histological examination of key developmental stages. Plant Cell Rep 23:115–119
Soundar Raju C, Kathiravan K, Aslam A, Shajahan A (2013) An efficient regeneration system via somatic embryogenesis in mango ginger (Curcuma amada Roxb.). Plant Cell Tiss Organ Cult 112:387–393
Soundar Raju C, Aslam A, Kathiravan K, Palani P, Shajahan A (2014) Direct somatic embryogenesis and plant regeneration from leaf sheath explants of mango ginger (Curcum amada Roxb.). In Vitro Cell Dev Biol Plant. doi:10.1007/s11627-014-9653-x
Stasolla C, Yeung EC (2003) Recent advances in conifer somatic embryogenesis: improving somatic embryo quality. Plant Cell Tiss Organ Cult 74:15–35
Viu AFM, Viu MAO, Tavares AR, Vianello F, Lima GPP (2009) Endogenous and exogenous polyamines in the organogenesis in Curcuma longa L. Sci Hortic 121:501–504
Xu ZQ, Jia JF, Hu ZD (1997) Somatic embryogenesis in Sesamum indicum L cv. nigrum. J Plant Physiol 150:755–758
You XL, Tan X, Dai JL, Li YH, Choi YE (2012) Large-scale somatic embryogenesis and regeneration of Panax notoginseng. Plant Cell Tiss Organ Cult 108:333–338
Zapata EV, Morales GS, Lauzardo ANH, Bonfil BM, Tapia GT, Sánchez ADJ, Valle MVD, Aparicio AJ (2003) In vitro regeneration and acclimatization of plants of turmeric (Curcuma longa L.) in a hydroponic system. Biotech Apl 20:25–31
Zuraida AR, Izzati KFL, Nazreena OA, Radziah CMZC, Asyikin SGSN, Sreeraman S (2014) In vitro regeneration of Curcuma caesia plantlets from basal part and via somatic embryogenesis. Adv Biosci Biotechnol 5:363–372
Acknowledgments
Dr. A. Shajahan thanks the University Grants Commission, Govt. of India for financial support (F. No. 42-946/2013-SR). The authors thank Dr. P. Palani, Centre for advanced studies in Botany, University of Madras for his help on scanning electron microscopic (SEM) observations. They also thank DST, Govt. of India for providing facilities through DST-FIST program.
Author information
Authors and Affiliations
Corresponding author
Rights and permissions
About this article
Cite this article
Soundar Raju, C., Aslam, A. & Shajahan, A. High-efficiency direct somatic embryogenesis and plant regeneration from leaf base explants of turmeric (Curcuma longa L.). Plant Cell Tiss Organ Cult 122, 79–87 (2015). https://doi.org/10.1007/s11240-015-0751-1
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s11240-015-0751-1