Multiplex logic processing isothermal diagnostic assays for an evolving virus

Sanchita Bhadra; Miguel A. Saldaña; Hannah Grace Han; Grant L. Hughes; Andrew D. Ellington

Most importantly, existing iNAATs lack design considerations to counter target variability, a deficiency that may reduce sensitivity 47 and in extreme cases lead to assay failure. For example, RPA uses a recombinase enzyme to facilitate invasion of double stranded templates by primers, which are then extended by a strand displacing DNA polymerase resulting in signal . CC-BY-NC-ND 4.0 International license is made available under a The copyright holder for this preprint (which was not peer-reviewed) is the author/funder. It . https://doi.org/10.1101/424440 doi: bioRxiv preprint amplification. 6 Amplicon detection is typically achieved using fluorophore-labeled probes that are measured using fluorimetry or converted to a color readout using lateral flow assays. 6, 48 NEAR relies on DNA nicking and extension reactions using nicking enzymes and strand displacing DNA polymerase to achieve exponential amplification of target DNA or RNA. 49 The resulting amplicons are typically converted to a fluorescence signal using cleavage probes. Neither configuration would be particularly suitable for degenerate priming and / or strand exchange computations, as opposed to our internally redundant Boolean logic processing assay.