Abstract
Somatic mutation detection is a fundamental component of cancer genome research and of the molecular diagnosis of patients’ tumors. Traditionally, such efforts have focused on either DNA exome or whole genome sequencing; however, we recently have demonstrated that integrating multiple sequencing technologies provides increased statistical power to detect mutations, particularly in low-purity tumors upon the addition of RNA sequencing to DNA exome sequencing. The computational protocol described here enables an investigator to detect somatic mutations through integrating DNA and RNA sequencing from patient-matched tumor DNA, tumor RNA, and germline specimens via the open source software, UNCeqR.
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References
Roychowdhury S, Chinnaiyan AM (2016) Translating cancer genomes and transcriptomes for precision oncology. CA Cancer J Clin 66:75–88
Roychowdhury S, Iyer MK, Robinson DR, Lonigro RJ, Wu YM, Cao X, Kalyana-Sundaram S, Sam L, Balbin OA, Quist MJ et al (2011) Personalized oncology through integrative high-throughput sequencing: a pilot study. Sci Transl Med 3:111ra121
Clinical Lung Cancer Genome P, Network Genomic M (2013) A genomics-based classification of human lung tumors. Sci Transl Med 5:209ra153
Meyerson M, Gabriel S, Getz G (2010) Advances in understanding cancer genomes through second-generation sequencing. Nat Rev Genet 11:685–696
Wilkerson MD, Cabanski CR, Sun W, Hoadley KA, Walter V, Mose LE, Troester MA, Hammerman PS, Parker JS, Perou CM, Hayes DN (2014) Integrated RNA and DNA sequencing improves mutation detection in low purity tumors. Nucleic Acids Res 42:e107
Li H, Handsaker B, Wysoker A, Fennell T, Ruan J, Homer N, Marth G, Abecasis G, Durbin R (2009) The Sequence Alignment/Map format and SAMtools. Bioinformatics 25:2078–2079
Li H, Durbin R (2009) Fast and accurate short read alignment with Burrows-Wheeler transform. Bioinformatics 25:1754–1760
Wang K, Singh D, Zeng Z, Coleman SJ, Huang Y, Savich GL, He X, Mieczkowski P, Grimm SA, Perou CM et al (2010) MapSplice: accurate mapping of RNA-seq reads for splice junction discovery. Nucleic Acids Res 38:e178
Troester MA, Hoadley KA, D’Arcy M, Cherniack AD, Stewart C, Koboldt DC, Robertson AG, Mahurkar S, Shen H, Wilkerson MD et al (2016) DNA defects, epigenetics, and gene expression in cancer-adjacent breast: a study from The Cancer Genome Atlas. NPJ Breast Cancer 2:16007
Mertins P, Mani DR, Ruggles KV, Gillette MA, Clauser KR, Wang P, Wang X, Qiao JW, Cao S, Petralia F et al (2016) Proteogenomics connects somatic mutations to signalling in breast cancer. Nature 534:55–62
Liu W, Snell JM, Jeck WR, Hoadley KA, Wilkerson MD, Parker JS, Patel N, Mlombe YB, Mulima G, Liomba NG et al (2016) Subtyping sub-Saharan esophageal squamous cell carcinoma by comprehensive molecular analysis. JCI Insight 1(16):e88755
Fishbein L, Leshchiner I, Walter V, Danilova L, Robertson AG, Johnson AR, Lichtenberg TM, Murray BA, Ghayee HK, Else T et al (2017) Comprehensive molecular characterization of pheochromocytoma and paraganglioma. Cancer Cell 31:181–193
Ciriello G, Gatza ML, Beck AH, Wilkerson MD, Rhie SK, Pastore A, Zhang H, McLellan M, Yau C, Kandoth C et al (2015) Comprehensive molecular portraits of invasive lobular breast cancer. Cell 163:506–519
The Cancer Genome Atlas Research Network (2014) Comprehensive molecular profiling of lung adenocarcinoma. Nature 511:543–550
The Cancer Genome Atlas Research Network (2012) Comprehensive genomic characterization of squamous cell lung cancers. Nature 489:519–525
The Cancer Genome Atlas Research Network (2015) Genomic classification of cutaneous melanoma. Cell 161:1681–1696
The Cancer Genome Atlas Research Network (2015) Comprehensive genomic characterization of head and neck squamous cell carcinomas. Nature 517:576–582
Brady SW, McQuerry JA, Qiao Y, Piccolo SR, Shrestha G, Jenkins DF, Layer RM, Pedersen BS, Miller RH, Esch A et al (2017) Combating subclonal evolution of resistant cancer phenotypes. Nat Commun 8:1231
Field MG, Durante MA, Anbunathan H, Cai LZ, Decatur CL, Bowcock AM, Kurtenbach S, Harbour JW (2018) Punctuated evolution of canonical genomic aberrations in uveal melanoma. Nat Commun 9:116
Yee T (2008) The {VGAM} package. R News 8:28–39
Gene Annotation File. https://api.gdc.cancer.gov/v0/data/a0bb9765-3f03-485b-839d-7dce4a9bcfeb
Karolchik D, Barber GP, Casper J, Clawson H, Cline MS, Diekhans M, Dreszer TR, Fujita PA, Guruvadoo L, Haeussler M et al (2014) The UCSC genome browser database: 2014 update. Nucleic Acids Res 42:D764–D770
Zook JM, Chapman B, Wang J, Mittelman D, Hofmann O, Hide W, Salit M (2014) Integrating human sequence data sets provides a resource of benchmark SNP and indel genotype calls. Nat Biotechnol 32(3):246–251
Sherry ST, Ward MH, Kholodov M, Baker J, Phan L, Smigielski EM, Sirotkin K (2001) dbSNP: the NCBI database of genetic variation. Nucleic Acids Res 29:308–311
Lek M, Karczewski KJ, Minikel EV, Samocha KE, Banks E, Fennell T, O'Donnell-Luria AH, Ware JS, Hill AJ, Cummings BB et al (2016) Analysis of protein-coding genetic variation in 60,706 humans. Nature 536:285–291
Forbes SA, Beare D, Gunasekaran P, Leung K, Bindal N, Boutselakis H, Ding M, Bamford S, Cole C, Ward S et al (2015) COSMIC: exploring the world's knowledge of somatic mutations in human cancer. Nucleic Acids Res 43:D805–D811
Cabanski CR, Wilkerson MD, Soloway M, Parker JS, Liu J, Prins JF, Marron JS, Perou CM, Hayes DN (2013) BlackOPs: increasing confidence in variant detection through mappability filtering. Nucleic Acids Res 41:e178
Cibulskis K, Lawrence MS, Carter SL, Sivachenko A, Jaffe D, Sougnez C, Gabriel S, Meyerson M, Lander ES, Getz G (2013) Sensitive detection of somatic point mutations in impure and heterogeneous cancer samples. Nat Biotechnol 31:213–219
Pickrell JK, Gilad Y, Pritchard JK (2012) Comment on “Widespread RNA and DNA sequence differences in the human transcriptome”. Science 335:1302 author reply 1302
Guo Y, Li J, Li CI, Long J, Samuels DC, Shyr Y (2012) The effect of strand bias in Illumina short-read sequencing data. BMC Genomics 13:666
Jiang Y, Turinsky AL, Brudno M (2015) The missing indels: an estimate of indel variation in a human genome and analysis of factors that impede detection. Nucleic Acids Res 43:7217–7228
Mose LE, Wilkerson MD, Hayes DN, Perou CM, Parker JS (2014) ABRA: improved coding indel detection via assembly-based realignment. Bioinformatics 30:2813–2815
Stouffer SA, Suchman EA, DeVinney LC, Star SA, Williams RM (1949) Studies in social psychology in World War II. Princeton University Press, Princeton, NJ
Wang K, Li M, Hakonarson H (2010) ANNOVAR: functional annotation of genetic variants from high-throughput sequencing data. Nucleic Acids Res 38:e164
Obenchain V, Lawrence M, Carey V, Gogarten S, Shannon P, Morgan M (2014) VariantAnnotation: a Bioconductor package for exploration and annotation of genetic variants. Bioinformatics 30:2076–2078
Lawrence MS, Stojanov P, Polak P, Kryukov GV, Cibulskis K, Sivachenko A, Carter SL, Stewart C, Mermel CH, Roberts SA et al (2013) Mutational heterogeneity in cancer and the search for new cancer-associated genes. Nature 499:214–218
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Wilkerson, M.D. (2019). A Computational Protocol for Detecting Somatic Mutations by Integrating DNA and RNA Sequencing. In: Krasnitz, A. (eds) Cancer Bioinformatics. Methods in Molecular Biology, vol 1878. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-8868-6_6
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DOI: https://doi.org/10.1007/978-1-4939-8868-6_6
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