QIAseq Human Exome Kits

• 疾患関連の遺伝子変異を迅速に、効率的に、確実に検出
• 1 日で完了するライブラリー調製ワークフローで、スケールアップも容易
• どなたでも簡単に扱えるデータ解析ツールと変異解釈ツールの併用で、データ解析時間を大幅短縮

QIAseq Human Exome Kit のターゲットサイズは 33 Mb で、GC リッチな領域であっても優れたカバレッジ均一性を実現し、SNV、Indel、CNV を高感度に検出可能です（SNV、Indel 併せて > 98%の感度を実現します）。リードを最大限に活用できるため、最大 50% のシークエンシングのコストダウンを実現します。また、1 日で完了するライブラリー調製ワークフローと、フレキシブルなハイブリダイゼーション時間の調整により、従来法より作業時間を 30% 削減します（FFPE や cfDNA サンプルでも、所要時間はたった30 分です）。

さらに、NGS 解析ツールである QIAGEN CLC Genomics Workbench と変異の解釈ツール QIAGEN Clinical Insights（QCI）を組合せることで、解析結果の解釈における時間と負担を最大 80% 削減することを可能にします。

エクソーム濃縮に使用する QIAseq Human Exome Kits には、エクソームプローブ、磁性ビーズ、濃縮試薬および増幅試薬が含まれています。

エクソーム濃縮前に使用するライブラリー調製キットは、別途購入が必要です。 Unique Dual Index (UDI) 対応の QIAseq FX DNA Library Kits を使用することで、2.5 時間のワークフローで、最大 384 個のマルチプレックスが可能なライブラリー調製が可能です。また、 QIAseq cfDNA Library Kits もしくは QIAseq Ultralow Input Library Kits と組合せることで、リキットバイオプシーや微量サンプルにも対応します。

The QIAseq Human Exome is a 37 Mb panel that delivers exceptional coverage uniformity regardless of the GC composition of the targets, facilitating >99% base-level coverage of targets at ≥20x and driving the comprehensive and sensitive detection of variants (>98% combined sensitivity for SNVs and Indels) while minimizing drop-out. The QIAseq Human Exome provides:

• Maximized read utilization resulting in up to 50% reduction in sequencing costs
• A single-day, automation-compatible sample to sequencing workflow, at least 33% faster than other exome workflows
• Pre-optimized analysis and variant interpretation workflows that reduce the burden of interpreting the tens of thousands of variants that result from exome sequencing

CNV detection is readily enabled due to the exceptional coverage uniformity provided by the panel in combination with the pre-optimized QIAseq Human Exome workflow within QIAGEN CLC Genomics Workbench. This approach delivers additional information not readily achievable with other commercially available exome sequencing solutions.

Eliminate the data bottleneck and reduce overall turnaround time with rapid data analysis and variant interpretation

Go from FASTQ file to high-quality variant calls in only 60 minutes with the QIAseq Human Exome workflow in QIAGEN CLC Genomics Workbench – 57% faster than the conventional BWA – GATK pipelines. With pre-configured single exome or trio workflows, SNVs, Indels and CNVs are identified accurately without the need for time-consuming optimization of analysis and filtering parameters. VCF files generated by this workflow are readily formatted for downstream interpretation.

Variant interpretation is powered by QIAGEN Clinical Insight (QCI) for QIAseq. This proven platform has been used to analyze and interpret over 1.6M NGS samples to date, leveraging the superior structured content of the QIAGEN Knowledge Base, which contains >20M biological and disease-related findings. Furthermore, end users are provided access to high-quality variant- and disease-specific evidence for interpretation, facilitating a better understanding of the role of these variants in disease pathogenesis.

Exome sequencing has been widely adopted in the last 10 years as an efficient way of screening the genome for disease-associated mutations. By focusing reads on coding regions, which harbor >80% of disease-causing mutations, the probability of identifying variants associated with disease is increased. At the same time, the amount of sequencing required is reduced by 99% compared to whole genomes, significantly minimizing the cost of sequencing.

The method is built upon the hybrid capture approach for target enrichment where probes, complementary in sequence to its targets, are designed and function as baits to “fish out" its target from a library of DNA molecules. Once the probes are hybridized to their targets, these probe-target hybrids are then bound to streptavidin-coated magnetic beads through a covalent interaction with the biotin label on the probes. A magnet is then used to keep these beads with the probe-target hybrids on the side of the tube while the rest of the DNA that is not of interest, is washed away, reducing off-target effects. After washing away unbound DNA, the targets are then amplified and prepared for sequencing.

Once the pre-capture libraries are generated, the libraries are hybridized to the exome probes. After hybridization, probe-target hybrids are then bound to streptavidin-coated magnetic beads and washed twice for stringency. Exome-enriched libraries are then amplified and prepared for sequencing on Illumina systems. Resulting FASTQ are uploaded into the QIAseq Human Exome workflow within the QIAGEN CLC Genomics Workbench for filtering, read mapping and variant calling. VCF output is then uploaded into QIAGEN Clinical Insight for QIAseq, which enables a variant filtering cascade that facilitates prioritization of variants for evidence-based interpretation.

QIAseq Human Exome Kits provide a scalable single-day, automation-compatible workflow that is adaptable to your unique needs or requirements of your application. Compatible with high-quality DNA, as well as cfDNA and DNA from FFPE samples, exceptional coverage uniformity and high-performance variant calling are achieved with variable hybridization times.

See the QIAseq Human Exome Handbook for more details.

Explore the QIAseq Human Exome BED file.

The QIAseq Human Exome protocol leverages the QIAseq FX library preparation for generating highly complex, unbiased libraries, even from samples of lower quality, such as DNA derived from FFPE, in only 2.5 hours. Additionally, QIAseq Human Exome is also compatible with the QIAseq cfDNA library prep. Libraries can be generated from enzymatically fragmented genomic DNA, physically sheared genomic DNA, as well as from circulating cellfree DNA. Depending on the DNA input type, we recommend using the following library preparation kits: QIAseq FX DNA Library Kits, QIAseq Ultralow Input Library Kits, and QIAseq cfDNA Library Kits (see the QIAseq Human Exome Handbook for more information).

QIAseq Human Exome Kits use a hybridization capture-based target enrichment approach to specifically enrich exonic sequences of the human genome from indexed whole genome libraries. The flexible workflow allows simultaneous hybridization capture from up to 8 samples with as little as 200 ng input per library.

Widespread adoption of exome sequencing has fueled many different, more cost-effective approaches to disease-based research. QIAseq Human Exome Kits can be used in a variety of applications that utilize exome sequencing, such as:

• Disease gene identification for rare and inherited disorders
• Population genetics and carrier screening
• Study of human disease pathways and mechanisms
• Study of pathogenic infection mechanisms

Disease gene identification and mutation analysis for rare and inherited disorders

Whole exome sequencing for rare disorders can be undertaken either as single exomes (proband [affected]-only) or as trio sequencing. Though the proband-only approach is more cost-effective, this typically results in a much higher number of variants compared to trio sequencing where both the biological parents are also sequenced in order to rule out private mutations and Mendelian inconsistencies, thereby narrowing down the set of variants for further investigation.

For mutation analysis in inherited disorders, exome sequencing is also becoming a widely used approach. By adopting an exome as a large panel from which disease-based gene sets can be screened, laboratories end up saving on reagent costs. In addition, labs can implement more standardized workflows, especially for automation, which decreases the introduction of type II errors. Furthermore, with the rapidly growing body of knowledge around human disease, the need to constantly update panels is mitigated by having an exome as a base panel since the exome already accounts for all the coding regions in the human genome.

Population genetics and carrier screening

The study of the pathogenesis of rare and complex diseases is complicated by the contribution of rare alleles that may exist in specific populations and variants that have a wide spectrum of allele frequencies that vary by ethnicity. Exome sequencing has been a tool widely used to profile various populations for such variants in order to better understand their frequencies within these specific groups. This enables better quantification of carrier risks, especially for debilitating diseases, enabling couples to plan better and make informed decisions, and for understanding susceptibility to the more common disorders that affect overall quality of life and/ or impact lifetime healthcare costs.

Study of human disease pathways and mechanisms

The utility of exomes extends beyond just identifying the disease-causing variant. In the effort to understand the mechanisms that feed into and regulate the pathways that are directly implicated in disease progression, typically with the intent to target them for treatment, exomes have also become a useful tool in conjunction with transcriptome sequencing. By sequencing the exome, other variants that may be acting as modifiers can be identified, thereby highlighting candidate regulatory pathways.

Study of pathogenic infection mechanisms

Bacterial or viral infection mechanisms, as well as what makes people susceptible to infection, are important to understand in order to design preventive and therapeutic measures to counteract the spread of disease. Exome sequencing provides an approach for helping identify mutations within proteins, either expressed at the cell-surface or involved in other transport mechanisms, that may provide a path for the pathogen to enter and thrive within the human host environment. These types of studies are structured very similarly to population studies wherein results are correlated with metadata on the disease within the population in order to establish significant results.