Combined Targeted DNA Sequencing in Non-Small Cell Lung Cancer (NSCLC) Using UNCseq and NGScopy, and RNA Sequencing Using UNCqeR for the Detection of Genetic Aberrations in NSCLC

PLoS One. 2015 Jun 15;10(6):e0129280. doi: 10.1371/journal.pone.0129280. eCollection 2015.


The recent FDA approval of the MiSeqDx platform provides a unique opportunity to develop targeted next generation sequencing (NGS) panels for human disease, including cancer. We have developed a scalable, targeted panel-based assay termed UNCseq, which involves a NGS panel of over 200 cancer-associated genes and a standardized downstream bioinformatics pipeline for detection of single nucleotide variations (SNV) as well as small insertions and deletions (indel). In addition, we developed a novel algorithm, NGScopy, designed for samples with sparse sequencing coverage to detect large-scale copy number variations (CNV), similar to human SNP Array 6.0 as well as small-scale intragenic CNV. Overall, we applied this assay to 100 snap-frozen lung cancer specimens lacking same-patient germline DNA (07-0120 tissue cohort) and validated our results against Sanger sequencing, SNP Array, and our recently published integrated DNA-seq/RNA-seq assay, UNCqeR, where RNA-seq of same-patient tumor specimens confirmed SNV detected by DNA-seq, if RNA-seq coverage depth was adequate. In addition, we applied the UNCseq assay on an independent lung cancer tumor tissue collection with available same-patient germline DNA (11-1115 tissue cohort) and confirmed mutations using assays performed in a CLIA-certified laboratory. We conclude that UNCseq can identify SNV, indel, and CNV in tumor specimens lacking germline DNA in a cost-efficient fashion.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adult
  • Aged
  • Aged, 80 and over
  • Carcinoma, Non-Small-Cell Lung / genetics*
  • Carcinoma, Non-Small-Cell Lung / pathology
  • Computational Biology
  • DNA Copy Number Variations
  • Female
  • Genetic Association Studies
  • Genetic Variation*
  • Genomics* / methods
  • High-Throughput Nucleotide Sequencing
  • Humans
  • Lung Neoplasms / genetics*
  • Lung Neoplasms / pathology
  • Male
  • Middle Aged
  • Mutation
  • Neoplasm Grading
  • Neoplasm Staging
  • Proto-Oncogene Proteins p21(ras) / genetics
  • Reproducibility of Results


  • KRAS protein, human
  • Proto-Oncogene Proteins p21(ras)