Improved DNase-seq protocol facilitates high resolution mapping of DNase I hypersensitive sites in roots in Arabidopsis thaliana.
BACKGROUND
Identifying cis-regulatory elements is critical in understanding the direct and indirect regulatory mechanisms of gene expression. 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. While this method was originally developed for human cell lines, later adaptations made the processing of plant tissues possible. Challenges still remain in processing recalcitrant tissues that have low DNA content.


RESULTS
By removing steps requiring the use of gel agarose plugs in DNase-seq, we were able to significantly reduce the time required to perform the protocol by at least 2 days, while also making possible the processing of difficult plant tissues. We refer to this simplified protocol as DNase I SIM (for simplified in-nucleus method). We were able to successfully create DNase-seq libraries for both leaf and root tissues in Arabidopsis using DNase I SIM.


CONCLUSIONS
This protocol simplifies and facilitates generation of DNase-seq libraries from plant tissues for high resolution mapping of DNase I hypersensitive sites.
Department of Botany and Plant Pathology, Oregon State University, Corvallis, OR 97331 USA.
Jason S
JS
Cumbie
Department of Botany and Plant Pathology, Oregon State University, Corvallis, OR 97331 USA.|Center for Genome Research and Biocomputing, Oregon State University, 2082 Cordley Hall, Corvallis, OR 97331 USA.
Sergei A
SA
Filichkin
Department of Botany and Plant Pathology, Oregon State University, Corvallis, OR 97331 USA.|Department of Electrical Engineering and Computer Science, Oregon State University, Corvallis, OR 97331 USA.|Center for Genome Research and Biocomputing, Oregon State University, 2082 Cordley Hall, Corvallis, OR 97331 USA.
Molly
0000-0001-6793-6151
M
Megraw
Arabidopsis; DNase I hypersensitive sites; DNase-seq; Nuclei; Open chromatin; Roots