Format

Send to

Choose Destination
J Vis Exp. 2013 Oct 18;(80):e50710. doi: 10.3791/50710.

Detecting somatic genetic alterations in tumor specimens by exon capture and massively parallel sequencing.

Author information

1
Department of Pathology, Memorial Sloan-Kettering Cancer Center.

Abstract

Efforts to detect and investigate key oncogenic mutations have proven valuable to facilitate the appropriate treatment for cancer patients. The establishment of high-throughput, massively parallel "next-generation" sequencing has aided the discovery of many such mutations. To enhance the clinical and translational utility of this technology, platforms must be high-throughput, cost-effective, and compatible with formalin-fixed paraffin embedded (FFPE) tissue samples that may yield small amounts of degraded or damaged DNA. Here, we describe the preparation of barcoded and multiplexed DNA libraries followed by hybridization-based capture of targeted exons for the detection of cancer-associated mutations in fresh frozen and FFPE tumors by massively parallel sequencing. This method enables the identification of sequence mutations, copy number alterations, and select structural rearrangements involving all targeted genes. Targeted exon sequencing offers the benefits of high throughput, low cost, and deep sequence coverage, thus conferring high sensitivity for detecting low frequency mutations.

PMID:
24192750
PMCID:
PMC3947962
DOI:
10.3791/50710
[Indexed for MEDLINE]
Free PMC Article

Supplemental Content

Full text links

Icon for MyJove Corporation Icon for PubMed Central
Loading ...
Support Center