Ultrarapid detection of SARS-CoV-2 RNA using a reverse transcription-free exponential amplification reaction, RTF-EXPAR
Carter, Jake G Orueta Iturbe, Lorea Duprey, Jean-Louis H A Carter, Ian R Southern, Craig D Rana, Marium Whalley, Celina M Bosworth, Andrew Beggs, Andrew D Hicks, Matthew R
Carter, Jake G
Orueta Iturbe, Lorea
Duprey, Jean-Louis H A
Carter, Ian R
Southern, Craig D
Rana, Marium
Whalley, Celina M
Bosworth, Andrew
Beggs, Andrew D
Hicks, Matthew R
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2021-08-31
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Abstract
A rapid isothermal method for detecting severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the virus responsible for COVID-19, is reported. The procedure uses an unprecedented reverse transcription-free (RTF) approach for converting genomic RNA into DNA. This involves the formation of an RNA/DNA heteroduplex whose selective cleavage generates a short DNA trigger strand, which is then rapidly amplified using the exponential amplification reaction (EXPAR). Deploying the RNA-to-DNA conversion and amplification stages of the RTF-EXPAR assay in a single step results in the detection, via a fluorescence read-out, of single figure copy numbers per microliter of SARS-CoV-2 RNA in under 10 min. In direct three-way comparison studies, the assay has been found to be faster than both RT-qPCR and reverse transcription loop-mediated isothermal amplification (RT-LAMP), while being just as sensitive. The assay protocol involves the use of standard laboratory equipment and is readily adaptable for the detection of other RNA-based pathogens.
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Carter JG, Orueta Iturbe L, Duprey JHA, Carter IR, Southern CD, Rana M, Whalley CM, Bosworth A, Beggs AD, Hicks MR, Tucker JHR, Dafforn TR. Ultrarapid detection of SARS-CoV-2 RNA using a reverse transcription-free exponential amplification reaction, RTF-EXPAR. Proc Natl Acad Sci U S A. 2021 Aug 31;118(35):e2100347118. doi: 10.1073/pnas.2100347118
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