This case study focuses on providing a more in-depth view of how the Daydreamer™ software enables rapid PCR assay design. Clostridium difficile is a common HAI. Phenotypic testing for C. difficile generally involves looking for the presence of at least one of two unique toxins typically produced by the infection, commonly called Toxin A and Toxin B. Many molecular diagnostics also target DNA sequences from the two corresponding genes.
We show that given a data set comprised of 87 toxigenic C. difficile draft genomes, 23 non-toxigenic C. difficile draft genomes, and the genomes of 88 other members of the Clostridium genus, Daydreamer™ rapidly identifies the gene region associated with toxigenicity. Furthermore, it precisely places primer sequences suitable for common real-time PCR protocols at exactly the most conserved sequences. The result is that the user can select one or more pairs perfectly conserved and unique to all toxigenic C. difficile regardless of the precise toxin phenotype, yielding a simple and robust detection strategy.
In conclusion, Daydreamer™ enables an in silico assay design process from genomic sequences that is rapid, robust, and suitable to a wide range of molecular assay design problems. Daydreamer™ is capable of utilizing platform-specific constraints as an integral part of its assay design process, resulting in an optimized design that is targeted exactly to the chemistry and platform to be utilized for a diagnostic.
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