QIAseq DIRECT SARS-CoV-2 utilizes a streamlined, 4-hour workflow for enrichment and library prep of the SARS-CoV-2 virus genome.
cDNA synthesis and SARS-CoV-2 enrichment
The QIAseq DIRECT SARS-CoV-2 workflow begins with random-primed cDNA synthesis (no rRNA depletion or poly-A selection required). This reaction is flexible with regard to input RNA; 5 µl viral RNA input is required as a starting volume, regardless of viral titer. Following cDNA synthesis, multiplexed primer pools are used in a high-fidelity multiplex PCR reaction to prepare two pools of approximately 250 bp amplicons. The two enriched pools per sample are then combined into a single tube and purified using a QIAseq Bead cleanup step.
Library amplification and sample indexing
Following quantification and normalization, SARS-CoV-2 enriched samples are amplified and sample-indexed using a high-fidelity amplification reaction. During this reaction, Unique Dual Indices (UDIs) are added to the samples. UDIs effectively mitigate the risk of read mis-assignment due to index hopping. This is enabled by filtering mis-assigned reads during the demultiplexing of individual samples, thus generating highly accurate output data. For more information on QIAseq UDIs, please refer to "Appendix A: QIAseq DIRECT Unique Dual Indexes" in the QIAseq DIRECT SARS-CoV-2 Handbook.
QIAseq DIRECT SARS-CoV-2 libraries are compatible with Illumina NGS platforms including iSeq 100, MiniSeq, MiSeq, NextSeq 500/550, HiSeq 2500, HiSeq 3000/4000 and NovaSeq 6000.