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Sample GSM2090072 Query DataSets for GSM2090072
Status Public on Apr 26, 2016
Title WT_H3K27me3
Sample type SRA
 
Source name young seedlings
Organism Arabidopsis thaliana
Characteristics developmental stage: 10-day-old whole seedlings
ecotype: C24
genotype: wild type
antibody: H3K27 (Abcam, ab6002, lot GR218433-4)
Treatment protocol No treatment
Growth protocol Seeds were grown on MS medium plates supplemented with 20 g/L sucrose and 0.8% agar at 22°C under long-day (23 h) illumination
Extracted molecule genomic DNA
Extraction protocol Genomic DNAs were extracted from seedlings using the DNeasy Plant Mini Kit (QIAGEN);total RNAs extracted from seedlings using the RNeasy Plant Mini Kit (QIAGEN)
For ChIP-seq library, a 10-ng quantity of ChIP or input DNA was used to prepare a high-throughput sequencing library. End repair, dA-tailing, adapter ligation, and amplification were carried out using the NEBNext® ChIP-Seq Library Prep Master Mix Set for Illumina (E6240, NEB) according to the manufacturer’s protocol. Index primers used for each sample were as follows: ATCACG for WT-H3K27me3, CGATGT for WT-H3K4me3, TTAGGC for WT-H3K9me2, TGACCA for WT-H3, ACAGTG for WT-Input, GCCAAT for pold2-H3K27me3, CAGATC for pold2-H3K4me3, ACTTGA for pold2-H3K9me2, GATCAG for pold2-H3, and TAGCTT for pold2-Input.
For Bisulfite-seq, a 2-μg quantity of genomic DNAs was sonicated (20 cycles of 30 s on, 30 s off, at low intensity) into 200-bp fragments with a Bioruptor (Diagenode, USA), end repaired, 3’-dA-tailed and ligated to methylated adapters. After purification, the ligated DNA fragments were treated with the EZ Methylation-Gold Kit (Zymo Research). The eluted bisulfite-treated DNAs were amplified using barcoded primers for sequencing (CGCTGT for the wild type and ACAGTG for the pold2 mutant)
 
Library strategy ChIP-Seq
Library source genomic
Library selection ChIP
Instrument model Illumina NextSeq 500
 
Data processing For ChIP-seq analysis, reads were mapped to the Arabidopsis genome (TAIR10) using bowtie1 (version 0.12.9) allowing two nucleotide mismatches using following parameters: bowtie -q -k 1 -n 2 -l 36 --best -S -p 2. Mapped reads were de-duplicated and sorted by samtools (version 0.1.19-44428cd) Peaks were found using MACS software (version 1.4.2) using H3 as a background. For H3K27me3 peaks finding in the wild type as an example, the parameters were as follow: macs14 -t WT_K27ME3.sorted.bam –c WT_H3.sorted.bam –f BAM -g 1.30e+8 -n WT_K27ME3 --shiftsize 73 --pvalue 1e-5 --bw=300 --mfold=10,30
For RNA-seq process, paired-end reads were selected and aligned to the Arabidopsis reference genome (version: TAIR10), using TopHat (v2.1.1) and Cuffdiff (v2.1.1).
For Bisulfite-seq analysis, high-throughput sequencing of the BS-seq library was performed on the Illumina NextSeq 500 System with single-end 75-bp reads. The raw sequence data were demultiplexed and converted to fastq files using bcl2fastq2 Conversion Software (version v2.16.0) under default parameters. Mapping, deduplication, and methylation extraction were performed using bismark software (V0.7.4),of which mapping using bowtie2 (2.0.5) to TAIR10.
Genome_build: tair10
 
Submission date Mar 15, 2016
Last update date May 15, 2019
Contact name Jinkui Cheng
E-mail(s) chengjinkui@cau.edu.cn
Organization name China Agricultural University
Lab Zhizhong Gong
Street address No.2 Yuanmingyuan West Road
City Beijing
ZIP/Postal code 100193
Country China
 
Platform ID GPL19580
Series (1)
GSE79259 The second subunit of DNA-polymerase delta is required for genomic stability and epigenetic regulation
Relations
BioSample SAMN04557557
SRA SRX1634365

Supplementary file Size Download File type/resource
GSM2090072_WT_H3K27me3.tdf 19.8 Mb (ftp)(http) TDF
GSM2090072_WT_K27ME3_peaks.bed.gz 105.3 Kb (ftp)(http) BED
SRA Run SelectorHelp
Processed data provided as supplementary file
Raw data are available in SRA

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