In vitro propagation and Agrobacterium-mediated transformation of safflower (Carthamus tinctorius L.) using a bacterial mutated aroA gene

Abstract

Callus induction and in vitro plantlet regeneration systems following Agrobacterium-mediated transformation using a mutated bacterial aroA gene were obtained in two cultivars of safflower (Carthamus tinctorius L.). Mean comparison showed that the highest percentage of induced callus was occurred on MS medium containing (1 mg l-1 NAA + 1 mg l-1 BAP) in cv. Dincer (94.33%) and (0.5 mg l-1 NAA + 0.5 mg l-1 BAP) in hypocotyl explant (97%), respectively. In addition, the highest percentage of shoot regeneration also was achieved on a range of media supplemented with 0.1 mg l-1 NAA + 2 mg l-1 BAP from cotyledon explant of Dincer cultivar (35.1%). In respect of transformation efficiency, the highest percentage of putative regenerated shoots on selective medium containing 50 mg l-1 kanamycin was achieved in cv. Dincer and LBA4404 strain (20.61%). PCR analysis of sixteen Dincer plantlets for both strains confirmed that mutated aroA gene was amplified using specific primers (EPS1F/EPS2R) and (EPS2R/35SF) yielded fragments of 1300 bp and 1800 bp, respectively whereas no plantlets contained this gene in Sina cultivar. This study revealed that cotyledon explant of cv. Dincer of safflower have a good potential for direct shoot regeneration, and A. tumefaciens LBA4404 can be established a beneficial method for the transformation of this oilseed crop by mutated aroA gene.