Automated primer design for DNA-based detection of the emerging potato pathogen Dickeya dianthicola
Shaista Karim: Colorado State University
<div>Soft-rotting Enterobacteria (SRE) comprise a subset of bacterial plant pathogens including agronomically important genera <em>Pectobacterium</em> sp. and <em>Dickeya</em> sp. Several <em>Pectobacterium</em> and <em>Dickeya </em>species represent significant pathogens of potato, but identifying these species with PCR is made difficult by their close relatedness and frequent genetic exchange. Numerous SRE species continuously impact potato yields but the recent emergence of <em>Dickeya dianthicola</em>, an invasive pathogen from Europe, now challenges disease management and diagnostic procedures across North America. To address this concern, we developed an automated comparative primer design pipeline, Uniqprimer, and used it to design primers for DNA-based detection of <em>D. dianthicola</em>. Uniqprimer performed alignments of six <em>D. dianthicola</em> genomes to 74 non-target genomes, designed primers based on regions of divergence, and mapped the primer pairs back to the genomes to confirm specificity. To demonstrate the specificity and sensitivity of our primers, we performed conventional and quantitative PCR assays. All primers were evaluated and confirmed for specificity to <em>D. dianthicola</em> using diverse bacterial isolates and infected potato tissues. Our results demonstrate that specific and sensitive PCR assays designed using Uniqprimer can aid in diagnosis and disease management of the emerging potato pathogen <em>D. dianthicola</em>. Uniqprimer is being tested for public release in the Galaxy web platform.</div>
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