Next Generation Sequencing
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Targeting cancer pathways: Sensitive, comprehensive detection of genomic alterations using a custom NGS panel

Xie J, Lu X, et al. (2016) Capture-based next-generation sequencing reveals multiple actionable mutations in cancer patients failed in traditional testing. Mol Genet Genomic Med, 4(3):262–272.


Cancer is a category of diseases, each of them broadly characterized by the type of cells from which the cancer originates. Key features that all types of cancer share are that cancer cells divide out of control of normal cellular mechanisms and do not differentiate into normal, functional cells. Commonly, cancer is treated with powerful chemotherapy drugs and radiation therapies that target dividing cells. This approach may destroy cancer cells, but it also can have significant health impacts on patients when healthy, noncancerous dividing cells are also destroyed. Additionally, such treatment regimens may leave behind residual cancer cells that are more aggressive than the cancer cells eliminated by the therapy.

Newer cancer therapies that target specific cancer pathways have fewer serious side effects. However, to deliver the correct therapy, clinicians must be able to determine which pathway is involved in a particular cancer case. Current next-generation sequencing (NGS) technologies are ideal for screening hundreds or thousands of variants, making these methods appealing for assessing the substantial genetic diversity of cancers.


Using xGen® Lockdown® Probes, Xie et al. describe the development of a complete genomic cancer panel for NGS that enriches for 115 genes that are either diagnostic for cancer or known to impact the effectiveness of specific cancer therapies. The scientists also developed a bioinformatics pipeline for identification of indels, base substitutions, and copy number variation—mutations that can affect gene expression and contribute to cancer development.

Xie et al. tested the performance of their assay using 14 patient samples that included formalin-fixed, paraffin-embedded (FFPE) tumor tissue, blood, and pleural fluid. Blood samples were sequenced at 50–100X and tumor samples at 200–500X mean coverage.

The researchers also describe some of the challenges of working with cancer samples, including low DNA content from small biopsy tissues or from mixed tissue specimens containing both healthy and tumor cells. In addition, many tumor biopsies are preserved as FFPE samples, which frequently contain fragmented nucleic acids that have been degraded by such factors as crosslinking and low pH. The authors present their optimization and library preparation methods for these challenging FFPE samples that allowed them to generate usable libraries from as little as 25 ng of genomic DNA.


The authors' optimized cancer panel and bioinformatics analysis protocols provided good sequencing uniformity and significantly reduced PCR duplicates. They reported 77–88% on-target performance for the 115 genes represented in the target capture panel.

Xie et al. note that the continued discovery of new cancer genes and genes with therapeutic implications will make it necessary to continually update their panel. For this, they emphasize the flexibility of xGen Lockdown Probes and the ease with which they can add new, custom DNA probes as these discoveries are made.

Product focus: Target capture reagents from IDT

xGen® Lockdown® Probes

xGen Lockdown Probes are individually synthesized, quality controlled, and normalized hybridization probes that offer:

  • Sensitive detection of SNPs, indels, CNV, LOH, and translocations
  • Flexibility to augment existing panels or create completely customized panels
  • Quick delivery

xGen Lockdown Probes for diagnostic use are also available. Contact for more information.

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)

Achieve uniform coverage with hybridization and wash buffers that are optimized for target enrichment 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 Universal Blockers—TS Mix is designed for use with Illumina® barcoded adapters. This ready-to-use mix effectively blocks dual- or single-index adapters to reducing adapter participation during in-solution hybridization capture and significantly improve on-target performance.

Blocking oligos are also available for other sequencing platforms. Custom adapters can be manufactured for other barcodes or to meet the needs of users who require specific modifications or services to improve performance in unique applicatioons.

Discover more about xGen Blocking Oligos.

Additional reading

Get the most out of your NGS samples—Learn about expandable NGS target capture panels that enrich for mutated genes implicated in tumors and genes associated with inherited diseases. Both panels return consistent results with high reproducibility and deep uniform coverage.

Delivering comprehensive genomic profiling for clinical cancer care—Scientists at Foundation Medicine, Inc. are leading a transformation in cancer care by helping clinicians to select appropriate treatment options for each patient, informed by a thorough understanding of the molecular changes specific to their disease. Read about the use of xGen Lockdown Probes in their flagship FoundationOne® Test.

Gene panels vs. gene-by-gene analysis for assessing disease risk—Citation summary: The authors compared an NGS-based gene panel and traditional testing data for diagnostic use and disease risk assessment in hereditary breast and ovarian cancer. Read how xGen Lockdown Probes were able to rescue drop-out regions of SureSelect® probe panels (Agilent).

Review other DECODED Online newsletter articles on NGS applications.

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

© 2016 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

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