Studies on genetic transformation of Theobroma cacao L.: evaluation of different polyamines and antibiotics on somatic embryogenesis and the efficiency of uidA gene transfer by Agrobacterium tumefaciens

Abstract

In order to develop a more efficient genetic transformation system for cacao somatic embryos, the effects of polyamines and b-lactam antibiotics on somatic embryogenesis, hygromycin as selective agent, and different factors affecting uidA gene transfer have been evaluated. The polyamines putrescine, spermidine, and spermine significantly improved secondary somatic embryogenesis in cacao. Spermine at 1,000 lM provided the best responses, increasing 6.79 the percentage of embryogenic callus and 2.59 the average number of embryos per embryogenic callus. The b-lactam antibiotics timentin and meropenem, used for Agrobacterium tumefaciens counter-selection, had a nondetrimental effect on secondary somatic embryogenesis, depending on their concentration, whereas the commonly used b-lactam cefotaxime inhibited it, irrespective of the tested concentration. Hygromycin showed a strong inhibitory effect on secondary somatic embryogenesis of cacao, impairing completely the embryo production at 20 mg l -1 . Following the criterion of GUS activity, the best conditions for T-DNA transfer into cotyledon explants from primary somatic embryos of cacao were a sonication of the explants for 100 s, a 20-min incubation period in Agrobacterium solution, an Agrobacterium concentration of 1.0 (OD 600 ), and cocultivation of the explants on tobacco feeder layers. These findings will have important implications for studies on functional genomics of cacao.

Cocoa Agronomy, Quality, Nutritional, and Health Aspects

“…The suitability and cost efficiency of this technique are under investigation and will determine the prospects for commercial application. Silva et al (2009) refined the aforementioned transformation system through their evaluation of the effects of polyamines and antibiotics on somatic embryogenesis and the efficiency of uidA gene transfer by Agrobacterium tumefaciens in the genetic transformation in cacao. Downloaded by [University of North Texas] at 08:35 04 December 2014…”

Overexpression of a 10-deacetylbaccatin III-10 β-O-acetyltransferase gene leads to increased taxol yield in cells of Taxus chinensis

“…The development of transgenic Taxus cells needs an efficient genetic transformation method. The Agrobacterium-mediated transformation system has been commonly applied to engineer many plant species for desirable traits (Li et al 2008;Gelvin 2009;Raffeiner et al 2009;Silva et al 2009;Banyai et al 2010;Espasandin et al 2010;Ma et al 2010;Zia et al 2010). This system is favored over other methods, such as particle bombardment, due to the integration of low numbers of the transgene into the genome, the stability in the host cells, and the high transformation efficiency, etc.…”

The effects of β-lactam antibiotics and hygromycin B on de novo shoot organogenesis in apple cv. Golden Delicious

“…This negative effect of timentin might be the consequence of a disruption of hormonal balance in apple tissue due to the presence of ticarcillin, clavulanic acid or their breakdown products. The inhibitory effect of timentin was also reported in somatic embryogenesis of walnut [55] and cocoa [56].…”