McQ - An open-source multiplexed SARS-CoV-2 quantification platform

ABSTRACT

McQ is a SARS-CoV-2 quantification assay that couples early-stage barcoding with high-throughput sequencing to enable multiplexed processing of thousands of samples. McQ is based on homemade enzymes to enable low-cost testing of large sample pools, circumventing supply chain shortages. Implementation of cost-efficient high-throughput methods for detection of RNA viruses such as SARS-CoV-2 is a potent strategy to curb ongoing and future pandemics. Here we describe Multiplexed SARS-CoV-2 Quantification platform (McQ), an in-expensive scalable framework for SARS-CoV-2 quantification in saliva samples. McQ is based on the parallel sequencing of barcoded amplicons generated from SARS- CoV-2 genomic RNA. McQ uses indexed, target-specific reverse transcription (RT) to generate barcoded cDNA for amplifying viral- and human-specific regions. The barcoding system enables early sample pooling to reduce hands-on time and makes the ap-proach scalable to thousands of samples per sequencing run. Robust and accurate quantification of viral load is achieved by measuring the abundance of Unique Molecular Identifiers (UMIs) introduced during reverse transcription. The use of homemade reverse transcriptase and polymerase enzymes and non-proprietary buffers reduces RNA to library reagent costs to 92 cents/sample and circumvents potential supply chain short-ages. We demonstrate the ability of McQ to robustly quantify various levels of viral RNA in 838 clinical samples and accu-rately diagnose positive and negative control samples in a test-ing workflow entailing self-sampling and automated RNA ex-traction from saliva. The implementation of McQ is modular, scalable and could be extended to other pathogenic targets in future.