1. Multiplexed and Extraction-Free Amplification for Simplified SARS-CoV-2 RT-PCR Tests
- Author
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Rafael Rivera, Amy Steadman, Crissa Bennett, John T. Connelly, Corrie Ortega, Bernhard H. Weigl, Paras Jain, S. Timothy Motley, Samantha A. Byrnes, and Ryan Gallagher
- Subjects
2019-20 coronavirus outbreak ,Coronavirus disease 2019 (COVID-19) ,Computer science ,Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) ,Economic shortage ,Computational biology ,010402 general chemistry ,Multiplexing ,01 natural sciences ,Sensitivity and Specificity ,Article ,Analytical Chemistry ,COVID-19 Testing ,Humans ,Multiplex ,Chemistry ,SARS-CoV-2 ,010401 analytical chemistry ,RNA ,COVID-19 ,Virology ,0104 chemical sciences ,Reverse transcription polymerase chain reaction ,Real-time polymerase chain reaction ,Rapid onset ,RNA, Viral ,RNA extraction ,Sample collection ,Multiplex Polymerase Chain Reaction - Abstract
The rapid onset of the global COVID-19 pandemic has led to multiple challenges for accurately diagnosing the infection. One of the main bottlenecks for COVID-19 detection is reagent and material shortages for sample collection, preservation, and purification prior to testing. Currently, most authorized diagnostic tests require RNA extraction from patient samples and detection by reverse transcription polymerase chain reaction (RT-PCR). However, RNA purification is expensive, time consuming, and requires technical expertise to perform. Additionally, there have been reported shortages of the RNA purification kits needed for most tests. With these challenges in mind, we report on extraction-free amplification of SARS-CoV-2 RNA directly from patient samples. In addition, we have developed a multiplex RT-PCR using the CDC singleplex targets. This multiplex has a limit of detection of 2 copies/μL. We have demonstrated these improvements to the current diagnostic workflow, which reduce complexity and cost, minimize reagent usage, expedite time to results, and increase testing capacity.
- Published
- 2021