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Protocol

Twist Total Nucleic Acids Library Preparation Kit for Viral Pathogen Detection and Characterization: For use with the Twist Target Enrichment Workflow

Viral pathogens come in all four types of genetic material: ssRNA, dsRNA, ssDNA and dsDNA. To address this diversity, Twist developed a single workflow that can be used to generate high quality sequencing libraries from any of these four types in parallel, enabling downstream Next Generation Sequencing (NGS) applications, including metagenomics and target enrichment. The Twist Total Nucleic Acids Library Preparation Kit for Viral Pathogen Detection and Characterization decreases cost and turn around time by consolidating multiple library preparation methods into a simple one-size-fits-all protocol. The protocol begins with denaturation and reverse transcription using random primers, followed by second strand synthesis with DNA polymerase I and E. Coli DNA ligase. The resulting mixture of dsDNA and cDNA is then converted into Illumina TruSeq-compatible libraries using the Twist Library Preparation Kit via Enzymatic Fragmentation and Unique Dual Indices (UDI). Libraries created with this method are compatible with Twist Target Enrichment Protocols and customizable viral detection panels. Synthetic viral RNA or DNA controls can be spiked into samples to serve as positive controls and to assess sensitivity.