Plant Gene Regulatory Networks pp 141-154
DNase I SIM: A Simplified In-Nucleus Method for DNase I Hypersensitive Site Sequencing
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 wordsDNase-seq DNase I hypersensitive sites Open chromatin Arabidopsis Roots Nuclei
- 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
- 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
- 8.Zhang W, Jiang J (2015) Genome-wide mapping of DNase I hypersensitive sites in plants. Methods Mol Biol 1284:71–89.Google Scholar