Targeted Sequencing

Overview

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.


Hybridization 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 SeqCap® EZ HyperCap workflow, an integrated workflow solution that combines high-efficiency KAPA DNA Library Preparation Kits with a performance-optimized, hybridization-based SeqCap EZ capture protocol. 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. The resultant high complexity libraries then serve as the starting material for the SeqCap EZ enrichment protocol.


Amplification Method

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. 

Available Products by Workflow
Workflow Step Product
Benefits
Sample QC
KAPA hgDNA Quantification & QC Kits
  • Reliable quantification and quality assessment of low-input or FFPE samples to optimize library construction yields and workflows
Target Enrichment

Hybridization

SeqCap EZ Target Enrichment Kits

SeqCap Epi Target Enrichment Kits

SeqCap RNA Enrichment Kits

Amplification

HEAT-Seq Target Enrichment Kits

  • Ability to focus only on the regions of interest
  • Comprehensive coverage of the exome and eipgenome
  • No fragmentation or library preparation necessary for HEAT-Seq Target Enrichment Kits

Library Preparation

KAPA HyperPrep Kits

KAPA HTP/LTP Library Preparation Kits

KAPA HyperPlus Kits

 

  • High-quality library construction from FFPE and challenging samples in less than 3 hours
  • Low bias enzymatic fragmentation for construction of high-quality libraries from microbes with extreme genomic GC content
Library Amplification
KAPA Library Amplification Kit
  • Minimal amplification bias for improved sequencing coverage in WGS workflows that require library amplification
Library Quantification 
KAPA Library Quantification Kits
  • Accurate quantification of adapter-ligated molecules prior to sample pooling and cluster generation or template preparation for optimal sequencing results