Next Generation Sequencing
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Analyzing the exome—focus your NGS analysis with high-performance target capture

The use of next generation sequencing (NGS) in discovery and diagnostic applications is rapidly expanding. Improvements in methods and reagents, as well as decreasing costs for sequencing, are the main drivers of this trend. High-throughput instruments sequence a remarkably large number of bases per run, but each instrument has an upper limit of bases that can be accurately read. For example, at its upper limit, the HiSeq® 2500 Instrument (Illumina) can sequence the human genome—approximately 3 billion base pairs—for about 10 individuals, at 20–30X coverage. While this is sufficient for investigating germline variations responsible for inherited traits or diseases, other applications may require larger sample sets or greater read depths.

By using a target enrichment strategy, researchers can focus the read capacity of their instrument to either run more samples, obtain more sequence reads in a subset of the genome, or both. The benefits of target enrichment are particularly important for sequencing rare variants, where hundreds of targeted reads are required to find key mutation(s) against a large genetic background. Areas where high read depth is indispensable include cancer research and studies of immune receptor diversity.

Of note, the target capture step can influence the quality of the final data, with panels and probe sets from some manufacturers performing poorly in GC-rich regions. This results has led some investigators to argue that it is better to just perform whole genome sequencing (WGS). However, we have demonstrated a method using individually synthesized, quality checked, DNA target capture probes (xGen® Lockdown® Probes) that eliminates the negative influence of poor captures. In her presentation titled, Analyzing the exome—focusing your NGS analysis with high performance target capture, Dr Kristina Giorda uses data from the xGen Exome Research Panel to show examples of how this target capture method performs across a variety of metrics and how it compares to other available solutions. Learn ways to improve your NGS target capture by watching the recorded presentation below.


Product focus: Target capture reagents

xGen® Lockdown® Probes—target capture probe pools for NGS

xGen Lockdown Probes are pools of individually synthesized, quality controlled, and normalized hybridization probes. Use them to generate custom capture panels for targeted sequencing to enhance the performance of existing panels. xGen Predesigned Gene Capture Probe Pools are available for any human RefSeq coding gene. Select from predesigned and custom probes that offer:

  • Sensitive detection of SNPs, indels, CNV, LOH, and translocations
  • GMP compliance for clinical and diagnostics research
  • Flexibility to augment existing panels or create completely custom panels
  • Quick delivery

Discover more about xGen Lockdown Probes.


xGen Lockdown Panels

xGen Lockdown Panels are preconfigured, validated, and stocked pools of xGen Lockdown Probes for targeted next generation sequencing of defined gene families:

  • xGen Exome Research Panel
  • xGen Acute Myeloid Leukemia Panel
  • xGen Pan-Cancer Panel
  • xGen Inherited Diseases Panel
  • xGen Human ID Research Panel
  • xGen Human mtDNA Research Panel

Discover more about xGen Lockdown Panels.


xGen Lockdown Reagents—hybridization and wash kit

xGen Lockdown hybridization and wash buffer reagents have been optimized to deliver deep, even coverage of targets captured using xGen Lockdown Probes and Panels. A short, 4-hour hybridization protocol generates results quickly.

Discover more about xGen Lockdown Reagents.


xGen Blocking Oligos

xGen Blocking Oligos prevent adapter cross hybridization and minimize off-target capture, increasing specificity during targeted sequencing. Universal Blocking Oligos—TS Mix are designed for use with Illumina barcoded adapters. This ready-to-use mix effectively blocks dual- or single-index adapters to significantly improve on-target performance.

Also available are:

  • xGen Standard Blocking Oligos
  • xGen Custom Blocking Oligos
  • xGen Blocking Oligos for diagnostics—contact GMPinfo@idtdna.com for information
  • xGen Blocking Oligos for Ion Torrent adapters

Discover more about xGen Blocking Oligos.


Related reading

Target enrichment facilitates focused next generation sequencing—Understand the rationale and benefits of enriching subsets of the genome (target enrichment by hybrid capture) prior to sequencing. Use this strategy for genotyping, identifying splice variants and indels, and profiling genomic recombination events as well as viral and transposon integration sites.

How important are those NGS metrics?—Of the many metrics used in evaluating target capture data for NGS applications, read about which ones our researchers consider important for evaluating performance of target enrichment panels.

Generate consistent, reliable exome sequencing results—Spanning 39 Mb of the human genome, the xGen® Exome Research Panel was designed to provide uniform and specific coverage of the coding regions for 19,396 genes. You can easily and cost-effectively expand this panel to include specific non-coding target regions through addition of xGen® Lockdown® Probes.

NGS Target Capture Custom Panels—determining probe number and cost—Learn about experimental design options that will determine the number (and cost) of probes required for an NGS target capture panel.


Review other DECODED Online newsletter articles on NGS applications.


You can also browse our DECODED Online newsletter for additional application reviews, lab tips, and citation summaries to facilitate your research.


Author: Hans Packer, PhD, is a scientific writer at IDT.


© 2017 Integrated DNA Technologies. All rights reserved. Trademarks contained herein are the property of Integrated DNA Technologies, Inc. or their respective owners. For specific trademark and licensing information, see www.idtdna.com/trademarks.


xGen® Blocking Oligos

Adapter blocking oligos increase the number of on-target reads by preventing non-specific binding during hybridization.

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