Molecular Methods in Oncology: Genomic Analysis

  • Jason A. JarzembowskiEmail author
Part of the Molecular Pathology Library book series (MPLB)


Next-generation sequencing (NGS) and chromosomal microarrays represent major technological advances—elegant combinations of biochemistry and microfluidics and laser optics—used in brute force approaches that rely on advanced computing to assemble and align the data from thousands or millions of individual reactions. NGS solutions comprise a spectrum of methods to obtain DNA sequence data in a massively parallel and automated fashion, which are faster and more efficient than the linear sequencing approach of three decades ago. Microarrays allow the simultaneous interrogation of numerous probes or targets, using sequence complementarity testing to identify similarities or differences in the genome. NGS and array-based comparative genomic hybridization provide insights into the whole spectrum of DNA aberrations, from single base substitutions to large-scale chromosomal deletions, which in turn help physicians diagnose and treat human disease.


Comparative genomic hybridization Microarray Mini-sequencing Next-generation sequencing Sanger sequencing Sequencing by oligonucleotide ligation and detection (SOLiD) Single-molecule real-time (SMRT) sequencing 


  1. 1.
    Sanger F, Nicklen S, Coulson AR. DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci U S A. 1977;74(12):5463–7.CrossRefPubMedPubMedCentralGoogle Scholar
  2. 2.
    Sanger F, Air GM, Barrell BG, Brown NL, Coulson AR, Fiddes CA, Hutchison CA, Slocombe PM, Smith M. Nucleotide sequence of bacteriophage phi X174 DNA. Nature. 1977;265(5596):687–95.CrossRefPubMedGoogle Scholar
  3. 3.
    Lander ES, Linton LM, Birren B, Nusbaum C, Zody MC, Baldwin J, International Human Genome Sequencing Consortium, et al. Initial sequencing and analysis of the human genome. Nature. 2001;409(6822):860–921. Erratum in: Nature 2001 Jun 7;411(6838):720. Szustakowki, J [corrected to Szustakowski, J]. Nature 2001 Aug 2;412(6846):565.CrossRefPubMedGoogle Scholar
  4. 4.
    Prober JM, Trainor GL, Dam RJ, Hobbs FW, Robertson CW, Zagursky RJ, Cocuzza AJ, Jensen MA, Baumeister K. A system for rapid DNA sequencing with fluorescent chain-terminating dideoxynucleotides. Science. 1987;238(4825):336–41.CrossRefPubMedGoogle Scholar
  5. 5.
    Smith LM, Sanders JZ, Kaiser RJ, Hughes P, Dodd C, Connell CR, Heiner C, Kent SB, Hood LE. Fluorescence detection in automated DNA sequence analysis. Nature. 1986;321(6071):674–9.CrossRefPubMedGoogle Scholar
  6. 6.
    Mardis ER. DNA sequencing technologies: 2006-2016. Nat Protoc. 2017;12(2):213–8.CrossRefPubMedGoogle Scholar
  7. 7.
    Metzker ML. Emerging technologies in DNA sequencing. Genome Res. 2005;15(12):1767–76.CrossRefPubMedGoogle Scholar
  8. 8.
    Pettersson E, Lundeberg J, Ahmadian A. Generations of sequencing technologies. Genomics. 2009;93(2):105–11.CrossRefPubMedGoogle Scholar
  9. 9.
    Illumina. Illumina sequencing technology. 2010. Product literature online at
  10. 10.
    Shendure J, Ji H. Next-generation DNA sequencing. Nat Biotechnol. 2008;26(10):1135–45.CrossRefPubMedGoogle Scholar
  11. 11.
    Canard B, Sarfati RS. DNA polymerase fluorescent substrates with reversible 3′-tags. Gene. 1994;148(1):1–6.CrossRefPubMedGoogle Scholar
  12. 12.
    Ion Torrent. Exome sequencing using the Ion Proton system. 2012. Product literature online at
  13. 13.
    Zhu Z, Jenkins G, Zhang W, Zhang M, Guan Z, Yang CJ. Single-molecule emulsion PCR in microfluidic droplets. Anal Bioanal Chem. 2012;403(8):2127–43.CrossRefPubMedGoogle Scholar
  14. 14.
    Pachauri V, Ingebrandt S. Biologically sensitive field-effect transistors: from ISFETs to NanoFETs. Essays Biochem. 2016;60(1):81–90.CrossRefPubMedPubMedCentralGoogle Scholar
  15. 15.
    Nakano K, Shiroma A, Shimoji M, Tamotsu H, Ashimine N, Ohki S, et al. Advantages of genome sequencing by long-read sequencer using SMRT technology in medical area. Hum Cell. 2017;30(3):149–61.CrossRefPubMedPubMedCentralGoogle Scholar
  16. 16.
    Rhoads A, Au KF. PacBio sequencing and its applications. Genomics Proteomics Bioinformatics. 2015;13(5):278–89.CrossRefPubMedPubMedCentralGoogle Scholar
  17. 17.
    Applied Biosystems. SOLiD™ system accuracy with the exact call chemistry module. 2011. White paper available at
  18. 18.
    Huang YF, Chen SC, Chiang YS, Chen TH, Chiu KP. Palindromic sequence impedes sequencing-by-ligation mechanism. BMC Syst Biol. 2012;6(Suppl 2):S10.CrossRefPubMedPubMedCentralGoogle Scholar
  19. 19.
    Bentley DR, Balasubramanian S, Swerdlow HP, Smith GP, Milton J, Brown CG, et al. Accurate whole human genome sequencing using reversible terminator chemistry. Nature. 2008;456(7218):53–9.CrossRefPubMedPubMedCentralGoogle Scholar
  20. 20.
    Gan C, Love C, Beshay V, Macrae F, Fox S, Waring P, Taylor G. Applicability of next generation sequencing technology in microsatellite instability testing. Genes (Basel). 2015;6(1):46–59.CrossRefGoogle Scholar
  21. 21.
    Nowak JA, Yurgelun MB, Bruce JL, Rojas-Rudilla V, Hall DL, Shivdasani P, Garcia EP, Agoston AT, Srivastava A, Ogino S, Kuo FC, Lindeman NI, Dong F. Detection of mismatch repair deficiency and microsatellite instability in colorectal adenocarcinoma by targeted next-generation sequencing. J Mol Diagn. 2017;19(1):84–91.CrossRefPubMedPubMedCentralGoogle Scholar
  22. 22.
    Salipante SJ, Scroggins SM, Hampel HL, Turner EH, Pritchard CC. Microsatellite instability detection by next generation sequencing. Clin Chem. 2014;60(9):1192–9.CrossRefPubMedGoogle Scholar
  23. 23.
    Campesato LF, Barroso-Sousa R, Jimenez L, Correa BR, Sabbaga J, Hoff PM, Reis LF, Galante PA, Camargo AA. Comprehensive cancer-gene panels can be used to estimate mutational load and predict clinical benefit to PD-1 blockade in clinical practice. Oncotarget. 2015;6(33):34221–7.CrossRefPubMedPubMedCentralGoogle Scholar
  24. 24.
    Goodman AM, Kato S, Bazhenova L, Patel SP, Frampton GM, Miller V, Stephens PJ, Daniels GA, Kurzrock R. Tumor mutational burden as an independent predictor of response to immunotherapy in diverse cancers. Mol Cancer Ther. 2017;16(11):2598–608.CrossRefPubMedGoogle Scholar
  25. 25.
    Johnson DB, Frampton GM, Rioth MJ, Yusko E, Xu Y, Guo X, Ennis RC, Fabrizio D, Chalmers ZR, Greenbowe J, Ali SM, Balasubramanian S, Sun JX, He Y, Frederick DT, Puzanov I, Balko JM, Cates JM, Ross JS, Sanders C, Robins H, Shyr Y, Miller VA, Stephens PJ, Sullivan RJ, Sosman JA, Lovly CM. Targeted next generation sequencing identifies markers of response to PD-1 blockade. Cancer Immunol Res. 2016;4(11):959–67.CrossRefPubMedPubMedCentralGoogle Scholar
  26. 26.
    Rosenberg JE, Hoffman-Censits J, Powles T, van der Heijden MS, Balar AV, Necchi A, Dawson N, O’Donnell PH, Balmanoukian A, Loriot Y, Srinivas S, Retz MM, Grivas P, Joseph RW, Galsky MD, Fleming MT, Petrylak DP, Perez-Gracia JL, Burris HA, Castellano D, Canil C, Bellmunt J, Bajorin D, Nickles D, Bourgon R, Frampton GM, Cui N, Mariathasan S, Abidoye O, Fine GD, Dreicer R. Atezolizumab in patients with locally advanced and metastatic urothelial carcinoma who have progressed following treatment with platinum-based chemotherapy: a single-arm, multicentre, phase 2 trial. Lancet. 2016;387(10031):1909–20.CrossRefPubMedPubMedCentralGoogle Scholar
  27. 27.
    Gubin MM, Artyomov MN, Mardis ER, Schreiber RD. Tumor neoantigens: building a framework for personalized cancer immunotherapy. J Clin Invest. 2015;125(9):3413–21.CrossRefPubMedPubMedCentralGoogle Scholar
  28. 28.
    du Manoir S, Speicher MR, Joos S, Schröck E, Popp S, Döhner H, et al. Detection of complete and partial chromosome gains and losses by comparative genomic in situ hybridization. Hum Genet. 1993;90(6):590–610.CrossRefPubMedGoogle Scholar
  29. 29.
    Kallioniemi A, Kallioniemi OP, Sudar D, Rutovitz D, Gray JW, Waldman F, et al. Comparative genomic hybridization for molecular cytogenetic analysis of solid tumors. Science. 1992;258(5083):818–21.CrossRefPubMedGoogle Scholar
  30. 30.
    Pinkel D, Albertson DG. Array comparative genomic hybridization and its applications in cancer. Nat Genet. 2005;37(Suppl):S11–7.CrossRefPubMedGoogle Scholar
  31. 31.
    Bier FF, von Nickisch-Rosenegk M, Ehrentreich-Förster E, Reiss E, Henkel J, Strehlow R, et al. DNA microarrays. Adv Biochem Eng Biotechnol. 2008;109:433–53.PubMedGoogle Scholar
  32. 32.
    Coughlin CR 2nd, Scharer GH, Shaikh TH. Clinical impact of copy number variation analysis using high-resolution microarray technologies: advantages, limitations and concerns. Genome Med. 2012;4(10):80.CrossRefPubMedPubMedCentralGoogle Scholar
  33. 33.
    Hacia JG, Collins FS. Mutational analysis using oligonucleotide microarrays. J Med Genet. 1999;36(10):730–6.CrossRefPubMedPubMedCentralGoogle Scholar
  34. 34.
    McGall GH, Christians FC. High-density genechip oligonucleotide probe arrays. Adv Biochem Eng Biotechnol. 2002;77:21–42.PubMedGoogle Scholar
  35. 35.
    Suomalainen A, Syvänen AC. Analysis of nucleotide sequence variations by solid-phase minisequencing. Methods Mol Biol. 2003;226:361–6.PubMedGoogle Scholar
  36. 36.
    Goto S, Takahashi A, Kamisango K, Matsubara K. Single-nucleotide polymorphism analysis by hybridization protection assay on solid support. Anal Biochem. 2002;307(1):25–32.CrossRefPubMedGoogle Scholar
  37. 37.
    Palmisano GL, Delfino L, Fiore M, Longo A, Ferrara GB. Single nucleotide polymorphisms detection based on DNA microarray technology: HLA as a model. Autoimmun Rev. 2005;4(8):510–4.CrossRefPubMedGoogle Scholar
  38. 38.
    Watson MA. Microarrays. In: Pfeifer JD, editor. Molecular genetic testing in surgical pathology. Philadelphia: Lippincott Williams & Wilkins; 2006.Google Scholar

Copyright information

© Springer International Publishing AG, part of Springer Nature 2018

Authors and Affiliations

  1. 1.Division of Pediatric PathologyMedical College of WisconsinMilwaukeeUSA

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