Transformation using RNAi technology for developing potato lines resistance to late blight (Phytophthora infestans)

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Phytophthora infestans (Mont.) de Bary causes late blight, a major disease of potato and tomato that causes production loss at about 10–100% in Indonesia. Breeding for late blight resistance in potato has been carried out in this country to minimize fungicide application. The objective of this study was to generate potato lines resistant to P. infestans through RNAi method. A series of studies were conducted, including confirmation of RNAi construct insertion in Agrobacterium tumefaciens and PCR analysis, transformation of two potato cultivars using A. tumefaciens carrying the RNAi fragment, in addition to selection and molecular analysis of selected plantlet using Polymerase Chain Reaction (PCR). The RNAi plasmid construct was confirmed via PCR analysis using specific primers for 35s and Tnos fragment, and resulted in 500 bp and 250 bp for fragment of 35s and Tnos, respectively. Transformation was performed on 733 Granola’s internode explants and 569 Atlantic’s internode explants. The transformation process produced 282 explants from Atlantic, while Granola did not produce any transformants. The level of transformation efficiency of Atlantic on selection medium containing hygromycin was 49.61%. Following regeneration step, the 282 selected explants produced 167 plantlets. Based on PCR reaction using specific primers for hpt gene, 14 plantlets were PCR positive and contained hpt fragment. Overall, Agrobacterium tumefaciens-mediated transformation on potato internode explants was successful. Therefore, the selected transformants should be further tested using bioassay for resistance to P. infestans.
Agrobacterium tumefaciens, Phytophthora infestans, RNAi, potato.