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QIAseq Targeted RNAscan Panelsは分子生物学的アプリケーション用であり、疾病の診断、予防、あるいは治療に使用することはできません。
This product contains substances regulated under REACH (EC 1907/2006 Annex XIV). The use of this product in the EU is permitted subject to an exemption (Article 56(3)). Please refer to the REACH notification and the SDS of this product, both of which can be found in the “Resources” section of this page, for more information.
特徴
- 分子バーコードによる融合遺伝子の正確な検出と定量解析
- ブレイクポイントの情報がなくても片側未知の融合遺伝子を同定可能
- FFPE やリキッドバイオプシーサンプルに対応
- わずか15 ng のRNA サンプルから解析可能
製品詳細
QIAseq Targeted RNAscan Panels は、既知および片側未知の融合遺伝子を定量的に解析することが可能なRNA シークエンス用ライブラリー調製試薬です。ターゲット遺伝子に分子バーコードでタグ付けしてからターゲット領域を増幅するため、PCR エラーと増幅バイアスの問題を克服できるユニークなアプローチです。また、独自のSingle Primer Extension（SPE）技術により、片側未知の融合遺伝子の検出が可能です。さらに、無償のデータ解析パイプラインGeneGlobe により、融合転写産物と新規融合遺伝子を高感度に検出することができます。
各QIAseq Targeted RNAscan Panel には、ターゲットエンリッチメントからライブラリー作製するために必要な試薬がすべて含まれています。本製品は、イルミナおよびサーモフィッシャーシークエンサーに対応しています。SPE技術によるプライマーデザインにより、各ターゲットは1つの遺伝子特異的プライマーと1つのユニバーサルプライマーによってターゲットエンリッチメントされるため、ターゲットエンリッチメントやライブラリー作製に必要なプライマーの種類およびプライマープールの数を削減することができます。イルミナおよびサーモフィッシャーシークエンサーに対応したサンプルインデックスを用いることで、シークエンシング1ランで最大384サンプルまでマルチプレックス可能です。
パフォーマンス
- Accuracy: Innovative digital sequencing (incorporating molecular barcodes) eliminates PCR duplication and amplification bias to confidently detect known and novel fusions.
- Specificity: The unique combination of our proprietary primer design algorithm and rigorous testing of every primer assay guarantees high specificity and accurate results.
- Content: The QIAseq targeted RNAscan panels offer a high degree of flexibility in content and sample multiplexing. Several cataloged panels have been developed for a wide range of applications. One can also build a custom panel for specific content, or to extend the contents of an existing cataloged panel. Up to 384 samples can be multiplexed using the QIAseq indexes.
- Flexibility: Because the QIAseq targeted RNAscan panels use single primer extension, primers can be designed to detect known fusions based on characterized breakpoints or to discover novel fusions.
原理
PCR duplicates are a major issue in targeted RNA sequencing for gene fusion detection, since – through PCR amplification – they turn unique RNA molecules into identical RNA molecules that cannot be distinguished from each other. This, in turn, results in the inability to confidently detect gene fusions. To overcome the issue of PCR duplicates, the QIAseq targeted RNAscan panels use digital sequencing by incorporating molecular barcodes into the starting RNA material before any amplification takes place – thereby preserving the uniqueness of the starting RNA molecules and overcoming the issues of not only PCR duplicates and amplification bias.
操作手順
The entire workflow of the QIAseq Targeted RNAscan Panels – from extracted RNA to sequencing-ready libraries – can be completed in 9 hours. Extracted RNA is converted to cDNA, targets are molecularly barcoded and enriched, and libraries are constructed. Sequencing files can be fed into the QIAseq pipeline – a cloud-based data analysis pipeline – which will filter, map and align reads, as well as count unique molecular barcodes associated with targeted regions and call fusions. This data can then be fed into QCI for interpretation.
アプリケーション
The QIAseq Targeted RNAscan Panels can be used to detect known and novel gene fusions from a wide range of sample types for numerous applications.
Sample types:
- FFPE
- Fresh or frozen tissue
- Cell lines
Applications:
- Detection of known gene fusions based on well-characterized breakpoints
- Discovery of novel gene fusions using exon- or gene-based primer designs
裏付けデータと数値
Workflow
Isolated RNA, as low as 20 ng, is converted to cDNA. This is followed by the library construction step, in which IL-N7 adapters, molecular barcodes, and sample indexes are incorporated into DNA fragments generated in the previous step. Library fragments now serve as templates for target enrichment using single primer extension. In this step, targets are enriched using a single gene-specific primer and a universal forward primer. The final step is library amplification and sample indexing (for dual indexing) using the IL-S5 sample index primer and a universal primer.
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