Agrobacterium mediated genetic transformati on of Papaver somniferum L. using semi solid agar gelled primed seeds as explant

S. Ozcan, K. M. Khawar, A. R. Osalou, S. D. Royandezagh

Abstract: The opium poppy Papaver somniferum is self-pollinating herbaceous annual plant with various rates of out-crossing and is cultivated since prehistoric times. It is used to obtain number of medicinally important alkaloids, rarely to obtain edible oil and is used as an ornamental plant. There is need to improve poppy by developing objective oriented superior cultivars. Traditional breeding methods for the development of improved new opium cultivars is very slow, due to high labor and lengthy breeding procedures. This makes it necessarry to look alternatives like developing easy and repeatable genetic transformation protocols. Previous reports in poppy suggest that it is very recalcitrant to in vitro regeneration, rooting and is difficult to acclimatize. Moreover, all regeneration protocols are variety and genotype dependent. The study aimed to develop an efficient genetic transformation protocol in poppy. To achieve the objective various explants including hypocotyl-cotyledon explant and poppy cultivar Ofis 96 seeds primed on agar solidified MS medium for 7 days were treated with with Agrobacterium tumefaciens strain GV 2260 harbouring p35 GUS INT plasmid for 30 minutes. Cocultivation was on MS medium for 48 hours. These seeds were selected on MS selection medium containing 50 mg/l Kanamycin and 500 mg/l Augmentin. After achieving length of 5-6 cm, the putative transgenic seedlings were transferred to pots for growth, development and acclimatisation. The leaves from acclimatised plants were subjected to GUS histochemical analysis. Gus positive plants were further confirmed by polymerase chain reaction (PCR). Putatve Transgenic plants were grown in glasshouse to flower and set seeds.

Keywords: Agrobacterium tumefaciens; GUS analysis; opium poppy; Papaver somniferum; PCR; transgenic

Date published: 2019-03-25