Comparison of High-Throughput Sequencing for Phage Display Peptide Screening on Two Commercially Available Platforms

  • Momoko TajiriEmail author


Phage display is a widely used technique to screen peptide sequences for interaction with target biomolecules, such as proteins and cells. Traditional protocols screen for only a limited fraction of a candidate pool due to the cost and time limitation of Sanger sequencing. Recent developments in high-throughput sequencing (HTS) technologies enable researchers to assess millions of biomolecule sequences. In this study, eluted DNA pooled from a phage display screening were sequenced by two types of HTS methodologies; sequence by synthesis and a nanopore-based techniques. While both methods resulted in the identification of several candidate peptide motifs determined to be interacting with the target proteins, the sequence by synthesis method provide a higher yield of valid reads for data analyses. Input library preparation protocol for the nanopore sequencer needs further modification to achieve effective data collection.


Peptide motifs Phage display Next-generation sequencing Bioinformatics Protein–protein interaction 



I would like to thank Michigan Technological University Research Excellence Fund (R01547) and Portage Health Foundation for funding (R75389).

Compliance with ethical standards

Conflict of interest

The author declares no conflict of interest.

Supplementary material

10989_2019_9858_MOESM1_ESM.docx (268 kb)
Supplementary material 1 (DOCX 268 kb)


  1. Bailey TL, Elkan C (1994) Fitting a mixture model by expectation maximization to discover motifs in biopolymers. Proc Int Conf Intell Syst Mol Biol 2:28–36PubMedGoogle Scholar
  2. Bailey TL, Machanick P (2012) Inferring direct DNA binding from ChIP-seq. Nucleic Acids Res 40(17):e128. CrossRefPubMedPubMedCentralGoogle Scholar
  3. Blankenberg D, Gordon A, Von Kuster G, Coraor N, Taylor J, Nekrutenko A, Galaxy T (2010) Manipulation of FASTQ data with Galaxy. Bioinformatics 26(14):1783–1785. CrossRefPubMedPubMedCentralGoogle Scholar
  4. Burley SK, Roeder RG (1996) Biochemistry and structural biology of transcription factor IID (TFIID). Annu Rev Biochem 65:769–799. CrossRefPubMedGoogle Scholar
  5. Dias-Neto E, Nunes DN, Giordano RJ, Sun J, Botz GH, Yang K, Arap W (2009) Next-generation phage display: integrating and comparing available molecular tools to enable cost-effective high-throughput analysis. PLoS ONE 4(12):e8338. CrossRefPubMedPubMedCentralGoogle Scholar
  6. Dinkel H, Van Roey K, Michael S, Kumar M, Uyar B, Altenberg B, Gibson TJ (2016) ELM 2016–data update and new functionality of the eukaryotic linear motif resource. Nucleic Acids Res 44(D1):D294–D300. CrossRefPubMedGoogle Scholar
  7. Feng Y, Zhang Y, Ying C, Wang D, Du C (2015) Nanopore-based fourth-generation DNA sequencing technology. Genomics Proteom Bioinform 13(1):4–16. CrossRefGoogle Scholar
  8. Filippakopoulos P, Knapp S (2012) The bromodomain interaction module. FEBS Lett 586(17):2692–2704. CrossRefPubMedGoogle Scholar
  9. Golparian D, Dona V, Sanchez-Buso L, Foerster S, Harris S, Endimiani A, Unemo M (2018) Antimicrobial resistance prediction and phylogenetic analysis of Neisseria gonorrhoeae isolates using the Oxford Nanopore MinION sequencer. Sci Rep 8(1):17596. CrossRefPubMedPubMedCentralGoogle Scholar
  10. Imai K, Tarumoto N, Misawa K, Runtuwene LR, Sakai J, Hayashida K, Maeda T (2017) A novel diagnostic method for malaria using loop-mediated isothermal amplification (LAMP) and MinION nanopore sequencer. BMC Infect Dis 17(1):621. CrossRefPubMedPubMedCentralGoogle Scholar
  11. Judge K, Harris SR, Reuter S, Parkhill J, Peacock SJ (2015) Early insights into the potential of the Oxford Nanopore MinION for the detection of antimicrobial resistance genes. J Antimicrob Chemother 70(10):2775–2778. CrossRefPubMedPubMedCentralGoogle Scholar
  12. Laver T, Harrison J, O’Neill PA, Moore K, Farbos A, Paszkiewicz K, Studholme DJ (2015) Assessing the performance of the Oxford Nanopore Technologies MinION. Biomol Detect Quantif 3:1–8. CrossRefPubMedPubMedCentralGoogle Scholar
  13. Lee M, Struhl K (2001) Multiple functions of the nonconserved N-terminal domain of yeast TATA-binding protein. Genetics 158(1):87–93PubMedPubMedCentralGoogle Scholar
  14. Ma Q, Zhang H, Mao X, Zhou C, Liu B, Chen X, Xu Y (2014) DMINDA: an integrated web server for DNA motif identification and analyses. Nucleic Acids Res 42((Web Server issue)):W12–W19. CrossRefPubMedPubMedCentralGoogle Scholar
  15. Machanick P, Bailey TL (2011) MEME-ChIP: motif analysis of large DNA datasets. Bioinformatics 27(12):1696–1697. CrossRefPubMedPubMedCentralGoogle Scholar
  16. Margos G, Hepner S, Mang C, Marosevic D, Reynolds SE, Krebs S, Fingerle V (2017) Lost in plasmids: next generation sequencing and the complex genome of the tick-borne pathogen Borrelia burgdorferi. BMC Genomics 18(1):422. CrossRefPubMedPubMedCentralGoogle Scholar
  17. Matochko WL, Chu K, Jin B, Lee SW, Whitesides GM, Derda R (2012) Deep sequencing analysis of phage libraries using Illumina platform. Methods 58(1):47–55. CrossRefPubMedGoogle Scholar
  18. Nikolov DB, Hu SH, Lin J, Gasch A, Hoffmann A, Horikoshi M, Burley SK (1992) Crystal structure of TFIID TATA-box binding protein. Nature 360(6399):40–46. CrossRefPubMedGoogle Scholar
  19. Sanchez R, Zhou MM (2009) The role of human bromodomains in chromatin biology and gene transcription. Curr Opin Drug Discov Dev 12(5):659–665Google Scholar
  20. Smith GP (1985) Filamentous fusion phage: novel expression vectors that display cloned antigens on the virion surface. Science 228(4705):1315–1317CrossRefPubMedGoogle Scholar
  21. Stafford RL, Zimmerman ES, Hallam TJ, Sato AK (2014) A general sequence processing and analysis program for protein engineering. J Chem Inf Model 54(10):3020–3032. CrossRefPubMedGoogle Scholar
  22. Tan Y, Tian T, Liu W, Zhu Z, J Yang C (2016) Advance in phage display technology for bioanalysis. Biotechnol J 11(6):732–745. CrossRefPubMedGoogle Scholar
  23. Thiel WH, Giangrande PH (2016) Analyzing HT-SELEX data with the Galaxy Project tools–A web based bioinformatics platform for biomedical research. Methods 97:3–10. CrossRefPubMedGoogle Scholar
  24. Thompson M (2009) Polybromo-1: the chromatin targeting subunit of the PBAF complex. Biochimie 91(3):309–319. CrossRefPubMedGoogle Scholar
  25. Weese D, Emde AK, Rausch T, Doring A, Reinert K (2009) RazerS–fast read mapping with sensitivity control. Genome Res 19(9):1646–1654. CrossRefPubMedPubMedCentralGoogle Scholar
  26. Wilson, B. D., Eisenstein, M., & Soh, H. T. (2019). High-Fidelity Nanopore Sequencing of Ultra-Short DNA Sequences. bioRxiv, 552224.
  27. Zhu X-Q, Li S-X, He H-J, Yuan Q-S (2005) On-column Refolding of an Insoluble His6-tagged Recombinant EC-SOD Overexpressed in Escherichia coli. Acta Biochim Biophys Sin 37(4):265–269. CrossRefPubMedGoogle Scholar

Copyright information

© Springer Nature B.V. 2019

Authors and Affiliations

  1. 1.Department of ChemistryMichigan Technological UniversityHoughtonUSA

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