SARS-CoV-2 detection with ApharSeq v1

• DOI: 10.17504/protocols.io.bif7kbrn
• Publication Date: 2020-07-11T08:54:25Z
• Authors: Daphna Strauss; Ayelet Rahat; Israa Sharkia; Alon Chappleboim; Miriam Adam; Daniel Kitsberg; Gavriel Fialkoff; Matan Lotem; Omer Gershon; Anna-Kristina Sch...; Esther Oiknine-Djian; Agnes Klochendler; Ronen Sadeh; Yuval Dor; Dana Wolf; Naomi Habib; Nir Friedman
• Keywords: Pooling; Amplicon

The global SARS-CoV-2 pandemic led to a steep increase in the need for viral detection tests worldwide. Most current are based on RNA extraction followed by quantitative reverse-transcription PCR assays that involve separate and qPCR reaction each sample with fixed cost time. While automation improved logistics can capacity of these tests, they cannot exceed this lower bound dictated one per sample. Multiplexed next generation sequencing (NGS) provide dramatic throughput, hold promise richer information strains host immune response. Here, we establish significant improvement existing RNA-seq protocols. Our workflow, ApharSeq (Amplicon Pooling Hybridization And RNA-Seq), includes fast cheap capture step, is coupled barcoding individual samples, sample-pooling prior reverse transcription, massively parallel sequencing. Thus, only step performed before pooling hundreds barcoded samples subsequent steps further analysis. Considering improvements, our proposed workflow estimated reduce costs 10-50 fold, labor 5-100 automated liquid handling 5-10 reagent requirements 100-1000 fold compared methods.