P rep are Li b rary | S eq uence | A nal yze Data | Sup p ort AmpliSeq™ for Illumina Targeted Resequencing Solution A streamlined, scalable amplicon sequencing solution, producing high-confidence data from low-input DNA and RNA samples. Highlights l Accurate Data Across Various Sample Types Proven AmpliSeq chemistry and Illumina sequencing technology combine to achieve high data quality, even from low-quality starting materials, such as FFPE tissues Comprehensive Content Portfolio Extensive menu of tested ready-to-use content and options for custom designs Fast, Simple Workflow Increased lab efficiency with a quick library preparation workflow needing < 1.5 hours of hands-on time Scalable Solution Multiple options for sequencing platforms, data analysis, and support meet virtually all throughput needs Introduction AmpliSeq for Illumina offers fast, accurate targeted resequencing that enables researchers to go from DNA or RNA to variant calls in 2.5 days. The supported solution includes curated panel content, a PCRbased library preparation assay, proven Illumina next-generation sequencing (NGS) technology, and user-friendly data analysis (Figure 1, Table 1). Delivered results provide the high on-target and uniform coverage needed to detect low-frequency variants with consistency and reliability. Leveraging high-performance AmpliSeq chemistry, researchers can use the AmpliSeq for Illumina assay to focus their studies on specific genes, regions, or variants of interest with high accuracy. Users can take advantage of expertly selected content in ready-to-use panels or create custom panels to meet specific needs. Custom options include on-demand gene panels for human disease research, preconfigured community panels, and custom DNA panels to target unique areas of interest. The PCR-based library preparation assay uses oligonucleotides to amplify regions of interest to investigate single nucleotide variants (SNVs), insert/deletions (indels), copy number variations (CNVs), and gene fusions. AmpliSeq chemistry can multiplex 12 to > 24,000 amplicons, enabling simultaneous capture of multiple targets in a single reaction. AmpliSeq for Illumina works with RNA and DNA samples, requiring as little as 1 ng of highquality DNA or RNA and the flexibility to accommodate formalin-fixed, paraffin-embedded (FFPE) samples, such as preserved tumor tissue. The AmpliSeq for Illumina solution is optimized for use with Illumina sequencing by synthesis (SBS) chemistry. Prepared libraries can be sequenced on any Illumina sequencing system, including the iSeq™ 100, MiniSeq™, MiSeq™, and NextSeq™ Sequencing Systems. Figure 1: AmpliSeq for Illumina Targeted Resequencing Solution For Research Use Only. Not for use in diagnostic procedures.

P rep are Li b rary | S eq uence | A nal yze Data | Sup p ort Table 1: AmpliSeq for Illumina At A Glance Parameter PCR-based workflow Learn more about AmpliSeq for Illumina ready-to-use panels at www.illumina.com/products/bybrand/ampliseq/ready-to-use-panels.html Sample Type FFPE tissue, blood Input Type Input Amount 1–100 ng; recommended 10 ng per pool Variant Types SNVs, indels, CNVs, gene fusionsa Hands-On Time (assay) Total Time (assay) 5–7.5 hours (panel dependent) Options for Content Ready-toUse Panel Predesigned panels targeting specific areas of interest Community Panel Designs...

P rep are Li b rary | S eq uence | A nal yze Data | Sup p ort Amplicon Assay and Library Preparation AmpliSeq for Illumina library preparation is fast and simple. A multiplexed, highly specific, PCR-based workflow results in libraries with high on-target alignment and high coverage uniformity (Figure 2). Up to 24,000 amplicons can be multiplexed in a single assay, allowing for simultaneous assessment of multiple genes. Library prep starts with amplifying specific regions in the starting DNA or cDNA . Remaining primer sequences are digested and sequencing adapters are added to the resulting...

a. Based on a panel with 1000 amplicons and a lOOOx mean coverage Figure 3: Illumina Sequencing Systems—Solutionsfor every application, sample type, and sequencing scale. Learn more at www.illumina.com/systems Figure 4: High Coverage Uniformity and On-Target Alignment in AmpliSeq for Illumina Custom Panels—Coriell and Horizon Discovery DNA samples were used to evaluate the performance of 11 AmpliSeq for Illumina custom panels of varying size (number of amplicons). Libraries were sequenced on the MiSeq System. Error bars indicate variability of technical replicates. High-Quality Data To demonstrate...