Polynucleotides for amplification and detection of SARS-CoV-2

Inventors

Andini, Nadya • Chiu, Kathy • Jiang, Xuewen • Maamar, Hédia

Assignees

Talis Biomedical Corp

Abstract

Disclosed herein are primers and probes related to the detection of SARS-CoV-2 via nucleic acid amplification testing (NAAT), for example to amplify and determine the presence of SARS-CoV-2 in test samples and/or to diagnose Covid-19. Specifically, the present disclosure describes primers and probes that bind to the N gene, ORF1ab, or E gene of SARS-CoV-2 coronavirus for detection via loop mediated isothermal amplification (LAMP) and molecular beacon hybridization.

Core Innovation

The disclosure describes SARS-CoV-2 NAAT using loop-mediated isothermal amplification (LAMP) with sequence-specific primer sets (Set-1 through Set-17). The primer sets target the viral N gene, ORF1ab, and E gene and are combined with labeled probes, including molecular beacons. The disclosure further specifies defined fluorophore/quencher pairs for the molecular beacons.

The disclosure recites RTLAMP detection workflows that include strand displacement DNA polymerase and reverse transcriptase. It further provides primer/probe sequence listing via SEQ ID-defined sets and molecular beacons, including the use of additional molecular beacon variants. The disclosure relates these sequence-specific components to SARS-CoV-2 NAAT performance.

Experimental examples report fast time-to-positive kinetics across primer sets, sensitivity down to tens of copies per reaction, and minimal cross-reactivity against common human coronaviruses. The disclosure also reports molecular-beacon based direct sequence discrimination and combined multi-target amplification using combinations of primer sets together with additional beacons to reduce false positives.

Claims Coverage

The partial content provides one independent claim, with dependent claims refining the composition by adding specific sequence elements and labeled detection components. Across the independent and dependent claims, there are multiple inventive features focused on a defined SEQ ID composition and optional probe/assay components.

Overall, the claim set coverage in the partial content centers on a composition defined by specific SEQ ID NO.s (63–68), with dependent refinements that add labeled probe components (molecular beacons with fluorophore/quencher and additional SEQ IDs, and an intercalating dye) and optionally add nucleic-acid enzymes (strand displacement polymerase and reverse transcriptase), as well as additional SEQ ID sets.

Stated Advantages

Documented Applications