GeneRead QIAact BRCA Advanced DNA UMI Panel

In verification studies conducted with FFPE tumor samples and high quality DNA extracted from Coriell immortalized tumor cell lines as starting material, the GeneRead QIAact BRCA 1/2 panel performed to a high level on the GeneReader NGS System. At a 5% variant allele frequency the workflow demonstrated 100% precision and sensitivity, successfully identifying all mutations present in the samples tested. A high degree of reproducibility was observed between sample batches, workflow operators and GeneReader systems.

DNA sequencing is a useful tool to detect genetic variations, including somatic mutations, single nucleotide variants (SNVs), copy number variation (CNVs) and insertions and deletions (inDels). Targeted enrichment technology enables next-generation sequencing (NGS) platform users to sequence specific regions of interest instead of the entire genome, effectively increasing sequencing depth and throughput with lower cost. Existing target enrichment methods, library preparation and sequencing steps all utilize enzymes and amplification processes, which introduce substantial bias and artifacts. These biases and artifacts lead to background artifactual errors that greatly limit the detection of true low-frequency variants in heterogeneous samples such as tumors. The GeneRead QIAact BRCA Advanced DNA UMIPanel integrates unique molecular index (UMI) technology into a gene-specific, primer-based target enrichment process, enabling sensitive variant detection of targeted genomic regions by NGS on the GeneReader system. The QIAact BRCA Advanced DNA UMI Panel has been optimized in combination with a specially formulated enrichment chemistry to achieve highly efficient enrichment on both regular and GC-rich regions at high multiplex levels.

The QIAact BRCA Advanced DNA UMI Panel is provided as four primer mix tubes, with up to 219 primers per tube. The QIAact BRCA Advanced DNA UMI Panel is comprised of two separate target enrichment panels, QIAact BRCA1/2 and QIAact pTEN/TP53, designed to enrich specific target regions in BRCA1, BRCA2 and pTEN, TP53 respectively. When used in combination, the QIAact BRCA Advanced DNA UMI Panel selectively targets all four genes. Genomic DNA samples are first fragmented, end-repaired and A-tailed using a single, controlled multi-enzyme reaction. The prepared DNA fragments are then ligated at their 5’ ends to a GeneReader specific adapter containing an UMI and a 9 base-pair (bp) sample-specific bar code. Ligated DNA molecules are subject to limited cycles of target enrichment PCR, with one gene-specific primer targeting a region and one universal forward primer complimentary to an adapter sequence. This reaction ensures that intended targets and UMIs are enriched sufficiently to be represented in the final library. A universal PCR with GeneReader specific sequences is then carried out to amplify the targets and complete the library. Once the library is sequenced, results can be analyzed using the GeneRead QIAact BRCA Advanced DNA UMI Panel QCI Analyze bioinformatics workflow, which will automatically perform all steps necessary to generate a DNA sequence variant report from your raw NGS data. All detected variants can be further interpreted by QIAGEN Clinical Insight (QCI), which is using QIAGEN’s comprehensive Knowledgebase of literature curated content to allow a standardized variant classification and reporting

For target enrichment prior to next-generation sequencing (NGS) applications that use the QIAGEN GeneReader instrument.Specifically for:

• Target enrichment and identification of somatic variants from FFPE samples
• Target enrichment and identification of germline variants from blood