Factors Affecting in Vitro Shoot Regeneration in Hypocotyls of Brinjal (Solanum melongena L.) in the Early Steps of Agrobacterium-Mediated Transformation
DOI:
https://doi.org/10.24154/jhs.v10i2.119Keywords:
Eggplant, Pre-Culture on High BAP, Explant Characters, PCRAbstract
An attempt was made to assess the effect of size, age and position of the explant, pre-culture and high cytokinin concentration in the pre-culture medium on shoot regeneration in brinjal hypocotyls co-cultivated with Agrobacterium. The study was carried out using hypocotyl explants of brinjal cv. Manjarigota, Agrobacterium strain A208 and shoot regeneration medium (full-strength basal MS medium, 2μM BAP + 0.05μM NAA, 3% sucrose and 0.8% agar) containing Cefotaxime (250-500mg l-1) and Kanamycin (100mg l-1). Hypocotyl explants showed callus initiation and shoot regeneration response after 10-12 and 20-22 days of culture, respectively. Five-day-old explants did not survive Agrobacterium infection, and ten-day-old explants showed higher shoot regeneration (29±1.91%) than older explants. Explants of medium size (1cm long; 32±2.62%) from the apical region (38.57±2.61%) showed better shootregeneration ability than explants of any other size or region. A period of four days of pre-culture (33.33±3.76) was optimal best for best shoot-regeneration in hypocotyl explants. No regeneration was seen in hypocotyl explants at shorter or longer pre-culture period. High cytokinin (10μM) in shoot regeneration medium during pre-culture enhanced shoot regeneration response (47.27±2.98%) in explants co-cultivated with Agrobacterium. Effects of various factors documented in this study will be useful in developing an efficient Agrobacterium-mediated transformation protocol in brinjal cv. Manjarigota.
Downloads
References
Balappa, S.R. and Hugar, L.B. 2002. An economic evaluation of brinjal production and its marketing system in Karnataka. Agril. Market, 44:45-49
Billings, S., Jelenkovic, G., Chin, C.K. and Eberhadt, J. 1997. The effect of growth regulation and antibiotics on eggplant transformation. J. Amer. Soc. Hortl. Sci., 122:158-162
Carman, J., Jefferson, N. and Cambell, W. 1988. Induction of embryogenic Triticum aestivum L. calli. II: Quantification of organic addenda and other culture variable effects. Pl. Cell Tiss. Org. Cult., 12:97-110
Chakrabarty, R., Viswakarma, N., Bhat, S.R., Kirti, P.B., Singh, B.D. and Chopra, V.L. 2002. Agrobacterium- mediated transformation of cauliflower: optimization of protocol and development of Bt-transgenic cauliflower. J. Biosci., 27:495-502
Chen, Q., Jelenkovic, G., Chin, C., Billings, S., Eberhardt, J. and Goffreda, J.C. 1995. Transfer and transcriptional expression of coleopteran Cry3B endotoxin gene of Bacillus thuringiensis in eggplant. J. Amer. Soc. Hort’l. Sci., 120:921-927
Chy, Y.S. and Philips, G.C. 1987. High efficiency Agrobacterium-mediated transformation of Lycopersicon based on conditions favaourable for regeneration. Pl. Cell Rep., 6:105-108
Collonier, C., Fock, I., Kashyap, V., Rotino, G.L., Daunay, M.C., Lian, Y., Mariska, I.K., Rajam, M.V., Servaes, A., Ducreux, G. and Sihachakr, D. 2001. Applications of biotechnology in eggplant. Pl. Cell Tiss. Org. Cult., 65:91-107
Compton, M.E. 2000. Interaction between explant size and cultivar affects shoot organogenic competence of watermelon cotyledons. Hort’l. sci., 35:749-750
Curuk, S., Elman, C., Schlarman, E., Sagee, O., Shomer, I., Cetiner, S., Gray, D.J. and Gaba, V. 2002. A novel pathway for rapid shoot regeneration from the proximal zone of the hypocotyl of melon (Cucumis melo L.). In Vitro Cell. Dev. Biol.-Plant, 38:260-267
Fari, M., Nagy, I., Csányi, M., Mityko, J. and Rásfalvy, A. 1995. Agrobacterium-mediated genetic transformation and plant regeneration via organogenesis and somatic embryogenesis from cotyledon leaves in eggplant (Solanum melongena L. cv. ‘Kecskemeti lila’). Pl. Cell Rep., 15:82-86
Frary, A. and Earle, E.D. 1996. An examination of factors affecting the efficiency of Agrobacterium-mediated transformation of tomato. Pl. Cell Rep., 16:235-240
George, E.F. 1993. Plant propagation by tissue culture. Part 1. The technology. Exegenetics Ltd., Eddington, England
Hamza, S. and Chupeau, Y. 1993. Re-evaluation of conditions for plant regeneration and Agrobacterium-mediated transformation from tomato. J. Expt’l. Bot., 44:1837-1845
Hanur, V.S., Prakash, D.P., Deepali, B.S., Asokan, R., Ramachandra, Y.L., Riaz Mahmood and Lalitha Anand. 2006. Synergistic use of hypocotyl explants and high BAP pre-conditioning for enhanced transformation frequency in brinjal (Solanum melongena L.). J. Hort’l. Sci., 1:116-119
Herath, P.S., Suzuki, T. and Hattori, K. 2005. Factors influencing Agrobacterium-mediated genetic transformation of Kenaf. Pl. Cell Tiss. Org. Cult., 82:201-206
Kumar, V.S. and Rajam, M.V. 2005. Enhanced induction of vir genes results in the improvement of Agrobacterium-mediated transformation of eggplant. J. Biochem. Biotech., 14:89-94
Lazzeri, P.A. and Dunwell, J.M. 1986. In vitro regeneration from seedling organs of Brassica oleracea var. italica Plenck. cv. Green Comet. II: Effect of light conditions and explant size. Annl. Bot., 58:699-710
Ling, H., Kriseleit, D. and Ganal, M.W. 1998. Effect of ticarcillin/ potassium clavulanate on callus growth and shoot regeneration in Agrobacterium-mediated transformation of tomato. Pl. Cell Rep., 17:843-847
Magioli, C. and Mansur, E. 2005. Eggplant (Solanum melongena L.): tissue culture, genetic transformation and use as an alternative model plant. Acta Bot. Brassilica, 19:139-148
Magioli, C., Rocha, A.P.M., Pinheiro, M.M., Martins, S.G. and Mansur, E. 2000. Establishment of an efficient Agrobacterium-mediated transformation system for eggplant and study of a potential biotechnologically useful promoter. J. Pl. Biotech., 2:43-49
Murashige, T. and Skoog, F. 1962. A revised method for rapid growth and bioassays with tissue cultures. Physiol. Pl., 15:473-497
Nehra, N.S., Chibbar, R.N., Kartha, K.K., Datla, R.S.S., Crosby, W.L. and Stushnoff, C. 1990. Agrobacterium-mediated transformation of strawberry calli and recovery of transgenic plants. Pl. Cell Rep., 9:10-13
Pastori, G.M., Wilkinson, M.D., Steeke, S.H., Sparks, C.A., Jones, H.D. and Parry, M.A.J. 2001. Age-dependent transformation frequency in wheat varieties. J. Expt’l. Bot., 52:857-63
Prakash, D.P., Deepali, B.S., Asokan, R., Ramachandra, Y.L., Lalitha Anand and Vageeshbabu S. Hanur. 2007a. Effect of explant type and growth regulators in the transformation of brinjal (Solanum melongena L.) cv. Manjarigota. J. Hort’l. Sci., 2:94-98
Prakash, D.P., Deepali, B.S., Asokan, R., Ramachandra, Y.L., Lalitha Anand and Vageeshbabu S. Hanur. 2007b. Effect of antibiotics and gelling agents in the transformation of brinjal (Solanum melongena L.) cv. Manjarigota. J. Hort’l. Sci., 2:19-25
Prakash, D.P., Deepali, B.S., Asokan, R., Ramachandra, Y.L., Shetti, D.L., Lalitha Anand and Vageeshbabu S. Hanur. 2008. Effect of growth regulators on in vitro complete plant regeneration in brinjal (Solanum melongena L.). Ind. J. Hort., 64:112-116
Sanyal, I., Singh, A.K., Kaushik, M. and Amla, D.V. 2005. Agrobacterium-mediated transformation of chickpea (Cicer arietinum L.) with Bacillus thuringiensis cry1Ac gene for resistance against pod borer insect Helicoverpa armigera. Pl. Sci., 168:1135-1146
Sharma, P. and Rajam, M.V. 1995. Genotype, explant and position effects on organogenesis and embryogenesis in eggplant (Solanum melongena L.). J. Expt’l. Bot., 46:135-141
Veluthambi, K., Gupta, K.A. and Sharma, A. 2003. The current status of plant transformation technologies. Curr. Sci., 84:368-379
Downloads
Published
Issue
Section
License
Copyright (c) 2015 D P Prakash, Y L Ramachandra, Vageeshbabu S Hanur (Author)
This work is licensed under a Creative Commons Attribution-NonCommercial-ShareAlike 4.0 International License.
Authors retain copyright. Articles published are made available as open access articles, distributed under the terms of the Creative Commons Attribution-NonCommercial-ShareAlike 4.0 International License, which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original author and source are credited. 
This journal permits and encourages authors to share their submitted versions (preprints), accepted versions (postprints) and/or published versions (publisher versions) freely under the CC BY-NC-SA 4.0 license while providing bibliographic details that credit, if applicable.
How to Cite
. 
