Rapid, versatile PCR

PROJECT SUMMARY The Polymerase Chain Reaction (PCR) is one of the most widely used methods in molecular diagnostics today. PCR amplifies short regions of DNA and, with a reverse transcription step to convert RNA to cDNA, PCR will amplify from RNA sequences. The importance of PCR was reinforced during the SARS-CoV- 2/COVID-19 pandemic, where its use as the de facto gold standard for monitoring infection made the acronym an everyday term in popular culture. PCR also became a standard method for monitoring SARS-CoV-2 in wastewater. While many advances in PCR methods and instrumentation have been made over the years, current instruments and methods involve multiple compromises in terms of speed, sample size, versatility, and portability. Notably, conventional benchtop PCR instruments (thermal cyclers) have a wide range of functionality including both endpoint (final product) PCR and real-time/quantitative PCR (qPCR), but they are slow, requiring 1-2 hours to obtain results. With this Phase I application, we propose advancement of a PCR instrument and methods that can achieve performance like a benchtop PCR instrument in terms of versatility, while also offering greatly improved speed, cost, and portability. This work builds on our prior experience on optical heating methods for PCR and takes advantage of recent technological advances to achieve the performance enhancements. Here we propose development of a rapid and versatile qPCR (rvPCR) instrument. Testing will be performed for detection of respiratory viruses, specifically SARS-CoV-2, Influenza A, and Influenza B.