A multiplexed, next generation sequencing platform for high-throughput detection of SARS-CoV-2

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Abstract

Population scale sweeps of viral pathogens, such as SARS-CoV-2, require high intensity testing for effective management. Here, we describe “Systematic Parallel Analysis of RNA coupled to Sequencing for Covid-19 screening” (C19-SPAR-Seq), a multiplexed, scalable, readily automated platform for SARS-CoV-2 detection that is capable of analyzing tens of thousands of patient samples in a single run. To address strict requirements for control of assay parameters and output demanded by clinical diagnostics, we employ a control-based Precision-Recall and Receiver Operator Characteristics (coPR) analysis to assign run-specific quality control metrics. C19-SPAR-Seq coupled to coPR on a trial cohort of several hundred patients performs with a specificity of 100% and sensitivity of 91% on samples with low viral loads, and a sensitivity of >95% on high viral loads associated with disease onset and peak transmissibility. This study establishes the feasibility of employing C19-SPAR-Seq for the large-scale monitoring of SARS-CoV-2 and other pathogens.

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  1. SciScore for 10.1101/2020.10.15.20212712: (What is this?)

    Please note, not all rigor criteria are appropriate for all manuscripts.

    Table 1: Rigor

    Institutional Review Board Statementnot detected.
    Randomizationnot detected.
    Blindingnot detected.
    Power Analysisnot detected.
    Sex as a biological variablenot detected.
    Cell Line Authenticationnot detected.

    Table 2: Resources

    Experimental Models: Cell Lines
    SentencesResources
    HEK293T RNA was extracted using the Total RNA extraction kit (Qiagen)
    HEK293T
    suggested: NCBI_Iran Cat# C498, RRID:CVCL_0063)
    Software and Algorithms
    SentencesResources
    All libraries were sequenced with MiSeq or NextSeq 500 (Illumina) using 75 bp paired-end sequencing.
    MiSeq
    suggested: (A5-miseq, RRID:SCR_012148)
    (scripts are available at https://github.com/seda-barutcu/FASTQstitch and https://github.com/seda-barutcu/MultiplexedPCR-DeepSequence-Analysis) The top enriched sequence variant from each sample is used for multiple alignment analysis using CLUSTALW.
    CLUSTALW
    suggested: (ClustalW, RRID:SCR_017277)

    Results from OddPub: Thank you for sharing your code.


    Results from LimitationRecognizer: An explicit section about the limitations of the techniques employed in this study was not found. We encourage authors to address study limitations.

    Results from TrialIdentifier: No clinical trial numbers were referenced.


    Results from Barzooka: We did not find any issues relating to the usage of bar graphs.


    Results from JetFighter: We did not find any issues relating to colormaps.


    Results from rtransparent:
    • Thank you for including a conflict of interest statement. Authors are encouraged to include this statement when submitting to a journal.
    • Thank you for including a funding statement. Authors are encouraged to include this statement when submitting to a journal.
    • No protocol registration statement was detected.

    About SciScore

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