Focuses sequencing efforts on the genomic regions of interest.
Targeted sequencing offers unique insights into specific regions of interest in the genome and is a powerful tool to investigate a variety of disease areas, including oncology, inherited diseases, immunology, infectious diseases, and more. Utilizing next-generation sequencing (NGS) technology, it allows targeting of specific genes, coding regions, even segments of chromosomes that are directly relevant, with precision and efficiency. Targeted sequencing is a more cost-effective method compared to whole genome sequencing (WGS) and aids in deeper analysis of results from WGS and other survey approaches.
Why targeted sequencing?
- Enables detection and quantification of rare and low-frequency variants
- Affords higher coverage, deeper sequencing and straightforward data analysis
- Provides cost effectiveness, time and resource savings, and precision
How does targeted sequencing work?
Target enrichment is a critical step in targeted sequencing. Specific probe sets are designed to enrich user-defined genomic regions of interest using either a hybridization or an amplification- based method.
In the hybrid capture method, pools of biotinylated oligonucleotide probes are designed to target specific regions of interest within a DNA fragment library. After a hybridization incubation, streptavidin-coated magnetic beads are used to attract the biotinylated probe/target hybrids. This results in a library highly enriched for the targeted DNA that is ready for sequencing.
Robust target enrichment and construction of libraries with maximum molecular complexity and minimal bias is critical for targeted sequencing applications. Roche Sequencing Solutions offers the HyperCap workflow, an integrated workflow solution that combines high-efficiency KAPA DNA Library Preparation Kits and KAPA (96) Dual-Indexed Adapters with performance-optimized, hybridization-based SeqCap EZ Probes and Roche® Universal Blocking Oligo (UBO) Kits. KAPA library preparation and amplification kits utilize optimally-formulated enzymes, including KAPA HiFi DNA Polymerase, evolved through our directed evolution technology. Protocols have been optimized to achieve superior library yields and limit amplification bias. KAPA Dual-Indexed Adapters enable high multiplexibility by utilizing up to 96 sample barcodes and UBO kits eliminate the need for adapter-matched blocking oligos, affording a simple and faster workflow. The resultant high complexity libraries then serve as the starting material for the SeqCap EZ enrichment protocol.
Current target enrichment methods via amplification rely on multiplex PCR or some form of primer extension across genomic regions of interest. The number of amplicons obtained can range from a single to thousands of targets, which can then be sequenced and analyzed. Typically, sequencing adapters are added to the amplicons via PCR in amplification methods. Thus a separate library construction is not required, which eases the workflow.
Roche Sequencing Solutions offers a portfolio of HEAT-Seq target enrichment products, which use validated molecular inversion probes (MIPS) for amplification-based target enrichment. They combine a fast and easy workflow with sensitive SNV detection and provide a sequencing-ready amplicon that contains the target DNA sequence, a molecular barcode, and sample indices.