Physical wounding-assisted Agrobacterium-mediated transformation of juvenile cotyledons of a biodiesel-producing plant, Jatropha curcas L.

Abstract

The non-edible plant Jatropha curcas L. is one of the most promising feedstock for sustainable biodiesel production as it is not a source of edible vegetable oils, produces high amounts of oil (approx. 30-60% in dry seeds) and does not require high-cost maintenance. However, as with other undomesticated crops, the cultivation of J. curcas presents several drawbacks, such as low productivity and susceptibility to pests. Hence, varietal improvement by genetic engineering is essential if J. curcas is to become a viable alternative source of biodiesel. There is to date no well-established and efficient transformation system for J. curcas. In this study, we tested various physical wounding treatments, such as sonication and sandvortexing, with the aim of developing an efficient Agrobacterium-mediated transformation for J. curcas. The highest stable transformation rate (53%) was achieved when explants were subjected to 1 min of sonication followed by 9 min of shaking in Agrobacterium suspension. The transformation frequency achieved using this protocol is the highest yet reported for J. curcas.

Establishment of bispyribac selection protocols for <i>Agrobacterium tumefaciens</i>- and <i>Agrobacterium rhizogenes</i>-mediated transformation of the oil seed plant <i>Jatropha curcas</i> L.

“…Li et al (2008) first reported the establishment of transgenic Jatropha from cotyledon explants by Agrobacterium tumefaciens-mediated transformation using the herbicide phosphinothricin as the selection agent. Recently, several groups have reported that transgenic Jatropha can be regenerated by hygromycin (Mao et al 2010;Kumar et al 2010b) and kanamycin selection (Pan et al 2010;Khemkladngoen et al 2011a). In most cases, there were substantial differences in the effects of plant growth regulators and optimized selection conditions, suggesting that there is large variation among di erent Jatropha genotypes and tissues in terms of their morphological responses to phytohormones.…”

Development of transgenic plants in jatropha with drought tolerance

“…Agrobacterium-mediated transformation was performed as described previously (Khemkladngoen et al 2011b), with some modifications. Physical wounding by sonication (1 min) followed by shaking (9 min) was applied to explants during Agrobacterium infection.…”

“…Physical wounding by sonication (1 min) followed by shaking (9 min) was applied to explants during Agrobacterium infection. After transformation, the cotyledonary explants were cultured and regenerated using the plant regeneration system as described by Khemkladngoen et al (2011b). e shoot regeneration medium was MS basal medium supplemented with 0.5 mg·l IBA was used as the root induction medium.…”

“…In this study, transformed explants were successfully regenerated using the regeneration system described by Khemkladngoen et al (2011b) with some modi cations for shoot regeneration and root induction. In addition, young leaves were used as plant materials.…”

“…us, an improved transformation method that includes wounding of tissue explants to facilitate the greater chances of Agrobacterium infection was established (Khemkladngoen et al 2011b). In the process of establishing a transformation method for jatropha, the regeneration of shoots from tissue explants also proved to be challenging.…”