DNase I SIM: A Simplified In-Nucleus Method for DNase I Hypersensitive Site Sequencing

Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1629)

Abstract

Identifying cis-regulatory elements is critical in understanding the direct and indirect interactions that occur within gene regulatory networks. Current approaches include DNase-seq, a technique that combines sensitivity to the nonspecific endonuclease DNase I with high-throughput sequencing to identify regions of regulatory DNA on a genome-wide scale. Yet, challenges still remain in processing recalcitrant tissues that have low DNA content. Here, we describe DNase I SIM (for Simplified In-nucleus Method), a protocol that simplifies and facilitates generation of DNase-seq libraries from plant tissues for high-resolution mapping of DNase I hypersensitive sites. By removing steps requiring the use of gel agarose plugs in DNase-seq, DNase I SIM reduces the time required to perform the protocol by at least 2 days, while also making possible the processing of difficult plant tissues including plant roots.

Key words

DNase-seq DNase I hypersensitive sites Open chromatin Arabidopsis Roots Nuclei 

References

  1. 1.
    Jiang J (2015) The ‘dark matter’ in the plant genomes: non-coding and unannotated DNA sequences associated with open chromatin. Curr Opin Plant Biol 24:17–23CrossRefPubMedGoogle Scholar
  2. 2.
    Song L, Crawford GE (2010) DNase-seq: a high-resolution technique for mapping active gene regulatory elements across the genome from mammalian cells. Cold Spring Harb Protoc 2010(2.) pdb prot5384.Google Scholar
  3. 3.
    John S, Sabo PJ, Canfield TK, Lee K, Vong S, Weaver M, Wang H, Vierstra J, Reynolds AP, Thurman RE, Stamatoyannopoulos JA (2013) Genome-scale mapping of DNase I hypersensitivity. Curr Protoc Mol Biol Chapter 27(Unit 21):27PubMedGoogle Scholar
  4. 4.
    Vierstra J, Wang H, John S, Sandstrom R, Stamatoyannopoulos JA (2014) Coupling transcription factor occupancy to nucleosome architecture with DNase-FLASH. Nat Methods 11(1):66–72CrossRefPubMedGoogle Scholar
  5. 5.
    Deal RB, Henikoff S (2011) The INTACT method for cell type-specific gene expression and chromatin profiling in Arabidopsis thaliana. Nat Protoc 6(1):56–68CrossRefPubMedGoogle Scholar
  6. 6.
    Sullivan AM, Arsovski AA, Lempe J, Bubb KL, Weirauch MT, Sabo PJ, Sandstrom R, Thurman RE, Neph S, Reynolds AP, Stergachis AB, Vernot B, Johnson AK, Haugen E, Sullivan ST, Thompson A, Neri FV 3rd, Weaver M, Diegel M, Mnaimneh S, Yang A, Hughes TR, Nemhauser JL, Queitsch C, Stamatoyannopoulos JA (2014) Mapping and dynamics of regulatory DNA and transcription factor networks in A. thaliana. Cell Rep 8(6):2015–2030CrossRefPubMedGoogle Scholar
  7. 7.
    Buenrostro JD, Giresi PG, Zaba LC, Chang HY, Greenleaf WJ (2013) Transposition of native chromatin for fast and sensitive epigenomic profiling of open chromatin, DNA-binding proteins and nucleosome position. Nat Methods 10(12):1213–1218CrossRefPubMedPubMedCentralGoogle Scholar
  8. 8.
    Zhang W, Jiang J (2015) Genome-wide mapping of DNase I hypersensitive sites in plants. Methods Mol Biol 1284:71–89.Google Scholar
  9. 9.
    Zhang W, Wu Y, Schnable JC, Zeng Z, Freeling M, Crawford GE, Jiang J (2012) High-resolution mapping of open chromatin in the rice genome. Genome Res 22(1):151–162CrossRefPubMedPubMedCentralGoogle Scholar
  10. 10.
    Zhang W, Zhang T, Wu Y, Jiang J (2012) Genome-wide identification of regulatory DNA elements and protein-binding footprints using signatures of open chromatin in Arabidopsis. Plant Cell 24(7):2719–2731CrossRefPubMedPubMedCentralGoogle Scholar
  11. 11.
    Cumbie JS, Filichkin SA, Megraw M (2015) Improved DNase-seq protocol facilitates high resolution mapping of DNase I hypersensitive sites in roots in Arabidopsis Thaliana. Plant Methods 11:42. doi:10.1186/s13007-015-0087-1 CrossRefPubMedPubMedCentralGoogle Scholar
  12. 12.
    Langmead B, Trapnell C, Pop M, Salzberg SL (2009) Ultrafast and memory-efficient alignment of short DNA sequences to the human genome. Genome Biol 10(3):R25CrossRefPubMedPubMedCentralGoogle Scholar
  13. 13.
    Boyle AP, Guinney J, Crawford GE, Furey TS (2008) F-Seq: a feature density estimator for high-throughput sequence tags. Bioinformatics 24(21):2537–2538CrossRefPubMedPubMedCentralGoogle Scholar

Copyright information

© Springer Science+Business Media LLC 2017

Authors and Affiliations

  1. 1.Department of Botany and Plant PathologyOregon State UniversityCorvallisUSA
  2. 2.Center for Genome Research and BiocomputingOregon State UniversityCorvallisUSA
  3. 3.Department of Electrical Engineering and Computer ScienceOregon State UniversityCorvallisUSA

Personalised recommendations