Efficient genetic transformation of chicory without selection marker
2015
Maroufi, A.
Description of the subject. The present study explains an Agrobacterium-mediated transformation method for chicory based on shoot regeneration medium containing no selective compounds (a non selection approach). Objectives. The aim of this experiment was to generate transgenic chicory plants on a shoot induction medium without selective compounds and study the inheritance of the integrated transgene. Method. Leaf explants were collected from in vitro grown seedlings of cultivar 'Melci'. Explants were inoculated with Agrobacterium harboring the pTJK136 plasmid. Callus induction and shoot regeneration were performed on the medium containing no antibiotics. Three separate transformation experiments were carried out. The presence of the transgene in regenerated plants (T0 and T1) was examined by PCR and RT-PCR. Furthermore, the regeneration frequency and transformation efficiency were separately calculated for each experiment. Results. Analytical results indicate that significant numbers of regenerated shoots recovered on a medium containing no selection agent are putative transformants. Screening of the regenerated plants from triplicate experiments revealed a remarkable transformation efficiency (up to 9%). Reverse transcription PCR (RT-PCR) analysis confirmed the correct expression of the transgene in the transformed plants and thus a stable integration in the genome. Additionally, to corroborate this last point, the transgene is successfully transmitted to the next generation (T1). Conclusions. Results confirmed the possibility of generating marker-free genetically modified plants in the form of a non-selection transformation system.
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