Direct shoot regeneration from leaf explants of Jatropha curcas in response to thidiazuron and high copper contents in the medium

Abstract

Leaf explants of Jatropha curcas cultured on Murashige and Skoog's (MS) medium supplemented with thidiazuron (TDZ; 0.90 µM) in combination with indole-3-butyric acid (IBA; 0.98µM) produced adventitious shoot buds directly on the surface of the explants without formation of intervening callus while shoot bud formation was accompanied with callus formation on medium supplemented with 6-benzylaminopurine (BAP; 13.3 µM) and IBA (2.46 µM). TDZ treatment resulted in more than twice higher rate of shoot bud induction than BAP. Shoot buds were multiplied and elongated following repeated transfers to medium containing BAP (2.22 µM) and gibberellic acid (GA 3 ; 1.44 µM). The effect of copper sulphate on differentiation of shoot buds from leaf segments was also investigated. Both shoot induction and multiplication media were supplemented with different levels of CuSO 4 (0 -5 µM). Significant improvement in shoot bud induction was observed when the concentration of CuSO 4 was increased to 10 times the normal MS level. Healthy elongated shoots were rooted on half strength MS medium supplemented with IBA (2.46 µM). Rooted plantlets were transferred to field and survived. Histological analysis revealed direct formation of shoot buds from leaf explants.Additional key words: adventitious bud regeneration, benzylaminopurine, gibberellic acid, indole-3-butyric acid, mineral nutrition. ⎯⎯⎯⎯The use of vegetable oil as biofuel is being considered as a suitable alternative to limited fossil fuel reserves. In this context, non-edible vegetable oil of Jatropha curcas has the potential of providing a promising and commercially viable alternative to petro-diesel (Gubitz et al. 1999). Jatropha curcas (family: Euphorbiaceae), is a multipurpose, drought resistant, perennial plant of Latin American origin, but it is now widespread throughout the tropical regions of the world (Openshaw 2000). Seeds are genetically heterozygous as Jatropha is cross pollinated, which results in a high degree of variation (Ginwal et al. 2005). To meet the large scale demand and ensure easy supply of the elite plant material, there is need to establish mass multiplication techniques. Though there are reports regarding the effect of different concentrations and combinations of plant growth regulators on regeneration from various explants in J. curcas Mukta 1996, Datta et al. 2007, Deore and Johnson 2008), attempts have not been made to standardize the nutrient requirement to improve regeneration. Primarily, the composition and ratios of plant growth regulators are manipulated to optimize the quality and number of organs initiated (Ramage and Williams 2002). However, a tissue growth rate and the extent and quality of morphogenic response are also influenced by the type and concentration of nutrients supplied in the culture medium (Niedz and Evens 2007). In the present investigation, we have developed an efficient regeneration protocol by standardizing the copper requirements for improved shoot bud induction.Young and fully-expanded leaves from apex were collected fr...

Efficient culture protocol for plant regeneration from petiole explants of physiologically mature trees ofJatropha curcasL.

“…In plant tissue culture, cytokinin is an essential factor for the induction of adventitious buds formation in most cases. [22,26,34À39] However, the mechanism underlying the induction of adventitious buds in our method is unclear. Our previous [33] and present studies suggest that our culture method might be applicable to a wide range of plant species, although different cytokinins should be tested for better results.…”

“…These results showed the genotypic effects on adventitious buds regeneration using the new culture method, which are in agreement with previous findings. [22,23] Effects of GA 3 on bud elongation GA 3 has beneficial effects on the elongation of regenerated adventitious buds in a woody plant Acacia mangium. [46] This effect is consistent with the role of GA 3 in stem elongation.…”

Analysis of organogenic competence of cotyledons of Jatropha curcas and their in vitro histological behavior

“…An important starting point in achieving this is the development of regeneration protocols suitable for mass propagation of the plant, to be used in molecular and genetic studies. Although there are a number of reports on the regeneration of the plant (Kalimuthu et al, 2007;Deore and Johnson, 2008;Li et al, 2008;Khurana-Kaul et al, 2010;Varshney and Johnson, 2010), further studies are still required in order to standardize, and in some cases validate, the different regeneration systems reported so far. For example, only recently has a report on histological investigations of the origin and developmental stages of morphogenesis of the plant in vitro appeared (Varshney et al, 2011).…”

High-frequency Adventitious Buds Induction from Leaf Explants of Jatropha Curcas L.

“…As a consequence, in order that the improved varieties can be cultivated via the genetic improvement method, it will be necessary to establish an efficient regeneration protocol properly to enhance the efficiency of adventitious buds regeneration and shorten the circle of tissue culture. By virtue of plant tissue cultures techniques, the regenerated plantlets have been obtained from different types of explants in J. curcas [10][11][12][13], but the regeneration effect of leaf explants is still remains to be improved and perfected expectantly [14][15][16].…”