Abstract
BackgroundOomycetes are pathogens of mammals, fish, insects and plants, and the potato late blight agent Phytophthora infestans and the oil palm and cocoa infecting pathogen Phytophthora palmivora cause economically impacting diseases on a wide range of crop plants. Increasing genomic and transcriptomic resources and recent advances in oomycete biology demand new strategies for genetic modification of oomycetes. Most oomycete transformation procedures rely on geneticin-based selection of transgenic strains.ResultsWe established N-acetyltransferase AAC(3)-I as a gentamicin-based selectable marker for oomycete transformation without interference with existing geneticin resistance. Strains carrying gentamicin resistance are fully infectious in plants. We further demonstrate the usefulness of this new antibiotic selection to super-transform well-characterized, already fluorescently-labelled P. palmivora strains and provide a comprehensive protocol for maintenance and zoospore electro-transformation of Phytophthora strains to aid in plant-pathogen research.ConclusionsN-acetyltransferase AAC(3)-I is functional in Phytophthora oomycetes. In addition, the substrate specificity of the AAC(3)-I enzyme allows for re-transformation of geneticin-resistant strains. Our findings and resources widen the possibilities to study oomycete cell biology and plant-oomycete interactions.
Highlights
Oomycetes are pathogens of mammals, fish, insects and plants, and the potato late blight agent Phytophthora infestans and the oil palm and cocoa infecting pathogen Phytophthora palmivora cause economically impacting diseases on a wide range of crop plants
We found that the neomycin phosphotransferase (nptII) selectable marker expressed by G418-resistant P. palmivora strains does not confer resistance to gentamicin, and that growth of P. palmivora strains carrying the aminoglycoside 3-N-acetyltransferase I (aacC1) selectable marker was arrested on V8 plates containing G418, but not gentamicin
In this study we highlight the usefulness of gentamicinbased selectable marker in oomycetes
Summary
Oomycetes are pathogens of mammals, fish, insects and plants, and the potato late blight agent Phytophthora infestans and the oil palm and cocoa infecting pathogen Phytophthora palmivora cause economically impacting diseases on a wide range of crop plants. The broad-host-range tropical species Phytophthora palmivora from clade 4 [3, 4] triggers disease on economically relevant crops including cocoa, mango, papaya, rubber tree, oil palm and many Citrus species [6, 7]. Phytophthora ramorum is threatening tanoak and other oak species in California and Oregon [8], and Phytophthora cinnamomi causes disease on multiple trees across the world, such as chestnut, oak, Eucalyptus and Banksia [9] Most of these species are spreading beyond their original geographic area due to international trade and climate change [2, 10]
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