|
| Status |
Public on Jul 22, 2015 |
| Title |
rosettes_wildtype_peg_H3_rep1 |
| Sample type |
SRA |
| |
|
| Source name |
rosette leaves
|
| Organism |
Arabidopsis thaliana |
| Characteristics |
tissue: rosette leaves
age: 21 days
chip antibody: H3
genotype: Col0
|
| Treatment protocol |
Control was kept in normal watered state, for other samples (peg) drought stress was induced by treatment with 30% Polyethylene glycol (PEG 6,000) for 5 hours.
|
| Growth protocol |
Columbia-0 and double mutant at3g03940/at518190 knockdown plants were grown in 12 hr light for 3 weeks in pots with soil covered with miracloth to prevent soil contamination of leaf tissues.
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Watered or water deficit treated vegetative tissues were fixed in 1% formaldehyde for 10 min after vacuum infiltration of the tissues in phosphate buffered saline (PBS) with 0.007% silwet. Fixation was stopped by quenching with 125 mM glycine. Next, the tissue was rinsed twice in ice cold water, blotted dry and flash frozen in liquid nitrogen. Chromatin was isolated and resuspended in 300 ml nuclei lysis buffer and sonicated for four 15 sec pulses using a Branson sonifier 450 at an output between 10 – 15%. 100 ml of sheared chromatin was diluted in 900 ml ChIP dilution buffer and precleared, using 5 ml of protein A Dynabeads (Invitrogen, Carlsbad, CA). Protein A beads were removed on a magnet, and the precleared chromatin was incubated overnight at 4 °C with antibodies against either mono-methylated H3K4 (10 mL, Abcam ab8895), dimethylated H3K4 (6 mL, Upstate 07-212), or trimethylated H3K4 (6 mL, Abcam ab8580). Next the antibody/chromatin complex was precipitated by incubating the mix with 40 ml protein A beads for 1 hour followed by collection of the beads on a magnet. The beads were washed and chromatin eluted and de-crosslinked using procedures. DNA was recovered by phenol/chloroform extraction and ethanol precipitation in the presence of Novagen pellet paint (CN Bioscience) and resuspended in 30 mL of TE buffer.
Purified DNA was end modified, ligated to amplification primers, amplified, and sequenced according to the manufacturer’s protocols (Illumina, San Diego, CA).
|
| |
|
| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina Genome Analyzer II |
| |
|
| Description |
Treated with 30% Polyethylene glycol (PEG 6,000) for 5 hours to induce drought stress
|
| Data processing |
Basecalls performed using CASAVA version 1.8.2
ChIP-seq reads were aligned to the Arabidopsis thaliana genome assembly (TAIR10) using bowtie version 0.12.8 with default parameters.
Tag density files where generated using the resulting SAM files from above step, and an in-house perl script.
Genome_build: TAIR10
Supplementary_files_format_and_content: Tab-delimited tag density.txt file: chromosome, start, end, count. Locations one-based.
|
| |
|
| Submission date |
Apr 28, 2015 |
| Last update date |
May 15, 2019 |
| Contact name |
Jean-Jack M. Riethoven |
| E-mail(s) |
jeanjack@unl.edu
|
| Organization name |
University of Nebraska-Lincoln
|
| Department |
Beadle Center for Biotechnology
|
| Lab |
Bioinformatics Core Research Facility
|
| Street address |
1901 Vine Street
|
| City |
Lincoln |
| State/province |
NE |
| ZIP/Postal code |
68588-0665 |
| Country |
USA |
| |
|
| Platform ID |
GPL9302 |
| Series (2) |
| GSE68370 |
Osmotic stress induces phosphorylation of histone H3 at threonine 3 in pericentromeric regions of Arabidopsis thaliana [ChIP-Seq] |
| GSE68440 |
Osmotic stress induces phosphorylation of histone H3 at threonine 3 in pericentromeric regions of Arabidopsis thaliana |
|
| Relations |
| BioSample |
SAMN03571037 |
| SRA |
SRX1012876 |
