Regeneration and transformation of purple passion fruit (Passiflora edulis S.) via Agrobacterium tumefaciens

Low fruit quality and short storage life caused by ethylene is a problematic issue in the passion fruit industry. Modification of plants with desirable traits by genetic engineering has emerged as an important route for eliminating the problem. The main objective of the present study was to find proper conditions for callus induction, plantlet regeneration and transformation of purple passion fruit with an antisense ACC oxidase gene mediated by Agrobacterium tumefaciens strain LBA4404 (pCAMBIA 1305.1). Calli could be successfully induced from hypocotyls of the passion fruit using MS solid medium supplemented with 0.5 mg L^-1 2,4-D combined with 1 mg L^-1 BA, resulting in the highest callus induction frequency of 100% and the callus size of 0.91 cm. MS solid medium supplemented with 3 mg L^-1 BA could be used as a regeneration medium for inducing callus development into whole plantlets, with the highest regeneration frequency of 83.3%. Genetic transformation of the passion fruit with the antisense ACC oxidase gene mediated by A. tumefaciens was successfully carried out by co-cultivating explants with Agrobacterium suspension for 15 min, followed by eliminating the bacterium with 300 mg L^-1 cefotaxime, and subsequently selecting transformed calli with 250 mg L^-1 kanamycin. Transformed calli were verified by histochemical GUS assay and the percentage of gus gene expression of 100% was observed. Molecular analysis by PCR confirmed the integration of transgene into plant genome.