AmpliSeq™ for Illumina BRCA Panel
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Prepare Library | Sequence | Analyze Data illumina* AmpliSeq™ for Illumina BRCA Panel Fast and accurate detection of somatic and germline mutations in BRCA1 and BRCA2. Highlights • Relevant Gene Content Target all exonic regions and flanking intronic sequences of BRCA1and BRCA2 • Fast, Streamlined Workflow Prepare sequencing-ready libraries in a single day using as little as 1 ng high-quality DNA or 10 ng DNAfrom FFPE tissue • Accurate Data Detect germline and somatic mutations down to 5% frequency using local orcloud-based analysis Table 1: AmpliSeq for Illumina BRCA Panel At A Glance Parameter Specification No. ofGenes 2 Introduction The AmpliSeq for Illumina BRCA Panel is a targeted resequencing assay designed for detecting somatic and germline mutations across all exonic regions and the flanking intronic sequences of BRCA 1 and BRCA2 (Table 1). BRCA1 and BRCA2 are human tumor suppressor genesthat, when carrying specific mutations, have been implicated in an increased riskfor breast and ovarian cancers.1 Understanding BRCA status within the tumor may be a factor when researching potential therapies.2 To assist with quick and accurate assessment of genomic variation within BRCA1 and BRCA2, Illumina offers the AmpliSeq for Illumina BRCA Panel. The BRCA panel is part of a streamlined workflowthat includes PCR-based library preparation, Illumina sequencing by synthesis (SBS) chemistry and next-generation sequencing (NGS) technology, and automated analysis. Requiring as little as 1 ng high-quality DNA per pool, the two-pool panel can be used with low-quality and low-quantity samples, including formalin-fixed, paraffin-embedded (FFPE) tissues. Simple, Streamlined Workflow The AmpliSeq for Illumina BRCA Panel is part of a DNA-to-variant solution that offers streamlined content, easy-to-perform library preparation, push-button sequencing systems, and simplified data analysis. Cumulative Target Size Variant Types Amplicon Size No. ofAmplicons Input DNA Requirement No. of Pools per Panel Supported Sample Types Germline-. Percent Targets Covered at Minimum 50x at Recommended Throughput Somatic. Percent Targets Covered at Minimum 500x at Recommended Throughput Coverage Uniformity (percent of targets with > 0.2x mean coverage) Percent On-Target Aligned Reads Total Assay Time Hands-On Time All exonic regionsofthe BRCA1 and BRCA2 tumor suppressor genes and flanking intronic sequences 22 kb SNVs, indelsa 98 bp on average 265 1-100 ng (10 ng recommended per pool) 2 FFPE tissue, blood DNA-to-Data Time 2.5 days a. SNVs. single nucleotide variations; indels. insertions/deletions b. Time represents library preparation only and does not include library quantification, normalization, or pooling Data on file at Illumina, Inc. 2017 Amplicon analysis workflow to perform alignment and variant calling. BaseSpace Sequence Hub provides access to BaseSpace Variant Interpreter, which assists with turning variant call data into annotated results. Learn more about Illumina informaticsatwww.illumina.com/ products/by-brand/ampliseq/ informatics.html Accurate Data Library preparation follows a straightforward, PCR-based protocol that can be completed in as little as 5 hours, with < 1.5 hours hands-on time. Resulting libraries can be normalized, pooled, and then loaded on to a flow cell for sequencing. Prepared libraries are sequenced using proven SBS chemistry on any compatible Illumina sequencing system (Table 2). Resulting data can be analyzed locally with Local Run Manageror easily streamed into BaseSpace™ Sequence Hub. Local Run Managerand BaseSpace Sequence Hub can access the DNA To demonstrate assay capabilities, Horizon Discovery (HD) and breast tissue (tumor/normal pair) samples were evaluated using the AmpliSeq for Illumina BRCA Panel on the MiSeq™ System. Results showed high coverage uniformity and on-target percentage of aligned reads, even with varying sample quality (Figure 1). In addition, two HD samples of varying quality were evaluated for variant calling accuracy. Data showed high concordance between expected and detected SNVs (Table 3). For Research Use Only. Not for use in diagnostic procedures 770-201 7-029-C | 1

Call Rate Table 2: Illumina Sequencing Systems Recommended for Use with the AmpliSeq for Illumina BRCA Panel No. of Samples per Run For investigating somatic mutations iSeq™ 100 System 12 17 hours MiniSeq™ System (mid output) 24 17 hours MiniSeq System (high output) 80 24 hours MiSeq System (v2 chemistry Nano) 3 17 hours MiSeq System (v2 chemistry Micro) 12 19 hours MiSeq System (v2 chemistry) 48 24 hours MiSeq System (v3 chemistry) 80 32 hours For investigating germline mutations iSeq™ 100 System 96 17 hours MiSeq System (v2...