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.
Passion fruit (Passiflora edulis (Sims)) is currently ranked third among fruit exports from Kenya and has great potential since the demand for both fresh fruit and processed juice is on a continuous increase. Passion fruit production in Kenya is constrained by a lack of healthy, clean planting material, poor seed viability, and low germination rates. To address this, the present study reports an in vitro plant regeneration protocol for passion fruit using leaf disc and nodal explants and genetic fidelity analysis of the regenerated plants. The highest number of shoot regeneration was obtained on Murashige and Skoog (MS) medium supplemented with 2 mg·L−1 6-Benzyl amino purine (BAP) (shoot induction medium). The multiplication of shoots was optimum in MS medium supplemented with 3 mg·L−1 BAP. To eliminate the requirement of an additional step of in vitro rooting, exogenous application of putrescine induced the formation and development of roots on nodal explants. Genetic fidelity analysis of the in vitro regenerated and macropropagated plants with that of the mother plant was carried out by sequence-related amplified polymorphism (SRAP) markers, and monomorphic banding profile for 80% of the regenerants confirmed the genetic uniformity of the in vitro regenerated and macropropagated plants. The in vitro regeneration system developed can be utilized for mass clonal propagation for the economic commercial exploitation of this important tropical fruit.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.