Plant Methods (Oct 2020)
A simple and fast Agrobacterium-mediated transformation system for passion fruit KPF4 (Passiflora edulis f. edulis × Passiflora edulis f. flavicarpa)
Abstract
Abstract Background Passion fruit (Passiflora edulis Sims) is an important horticultural crop in the tropics and subtropics, where it has great commercial potential due to high demand for fresh edible fruits and processed juice as well as source of raw materials in cosmetic industries. Genetic engineering shows great potential in passion fruit improvement and can compensate for the limitations of conventional breeding. Despite the success achieved in genetic modification of few passion fruit varieties, transgenic passion fruit production is still difficult for farmer-preferred cultivars. Therefore, it is important to establish a simple and fast Agrobacterium-mediated cell transformation of commercial hybrid passion fruit KPF4 (Passiflora edulis f. edulis × Passiflora edulis f. flavicarpa). Results In the present study, we have developed a simple and fast Agrobacterium-mediated transformation system for hybrid passion fruit KPF4 using leaf disc explants. Factors affecting the rate of transient beta (β)-glucuronidase (gusA) expression and consequently transformation efficiency were optimized as follows: Agrobacterium cell density with an OD600 of 0.5, 30 min infection time, 3 days of co-cultivation duration and the incorporation of 200 µM acetosyringone into Agrobacterium infection suspension medium. Using the optimized conditions, transgenic plants of KPF4 were produced within 2 months with an average transformation efficiency of 0.67%. The β-glucuronidase (GUS) histochemical staining confirmed the expression and integration of an intron-containing gusA gene into transformed leaf discs and transgenic plant lines of KPF4. The presence of gusA gene in the transgenic plants was confirmed by polymerase chain reaction (PCR). The results confirmed that the gusA gene was efficiently integrated into the passion fruit genome. Conclusions The developed transformation protocol is simple and rapid and could be useful for functional genomic studies and transferring agronomically important traits into passion fruit hybrid KPF4. This study developed a method that can be used to transfer traits such as resistance to viral diseases, low fruit quality and short storage life. To the best of our knowledge, this is the first report on genetic transformation system for commercial passion fruit hybrid KPF4.
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