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Sample GSM2101796 Query DataSets for GSM2101796
Status Public on May 28, 2016
Title TWT_H3
Sample type SRA
Source name young seedlings
Organism Arabidopsis thaliana
Characteristics developmental stage: 10-day old whole seedlings
ecotype: C24
genotype: transgenic wild type
chip-antibodies: H3 (Abcam, ab1791, lot GR177884-2)
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 For RNA-seq, total RNAs extracted from seedlings using the RNeasy Plant Mini Kit (QIAGEN). For ChIP-seq,A 2-g sample of the 10-day-old seedlings was crosslinked with 1% formaldehyde. Chromatin were isolated using nuclei extraction buffer (1 M hexylene glycol, 20 mM Tris-HCl (pH 8.0), 0.15 mM spermine, 5 mM 2-mercaptoethanol, 1% Triton X-100, 0.1 mM PMSF, and cocktail) and precipitated by centrifuging at 2,000 × g for 10 min at 4°C. After resuspension in 300 μL of nuclei lysis buffer (50 mM Tris-HCl (pH 8.0), 10 mM EDTA, 1% sodium dodecyl sulfate [SDS] and cocktail) at 4°C, the chromatin was sonicated into fragments of approximately 300 bp using a Bioruptor (15 cycles of 30 s on and 30 s off at high intensity)
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: AGTCAA for TWT-H3K27me3; GTCCGC for TWT-H3K4me3; AGTTCC for TWT-H3K9me2; ATGTCA for TWT-H3; CCGTCC for TWT-Input; GTGGCC for fen-H3K27me3; GTGAAA for fen-H3K4me3; GTTTCG for fen-H3K9me2; CGTACG for fen-H3; and GAGTGG for fen-Input.
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 TWT_H3.sorted.bam –f BAM -g 1.30e+8 -n WT_K27ME3 --shiftsize 73 --pvalue 1e-5 --bw=300 --mfold=10,30.TDF files were generated using igvtools count for IGV visualization.
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). The output accepted_hits.bam files were used to generate TDF files by igvtools count for IGV visualization.
Genome_build: tair10
Submission date Mar 30, 2016
Last update date May 15, 2019
Contact name Jinkui Cheng
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)
GSE79738 Requirement for flap endonuclease 1 (FEN1) to maintain genomic stability and transcriptional gene silencing in Arabidopsis
BioSample SAMN04591553
SRA SRX1670669

Supplementary file Size Download File type/resource
GSM2101796_TWT_H3.tdf 19.8 Mb (ftp)(http) TDF
SRA Run SelectorHelp
Raw data are available in SRA
Processed data provided as supplementary file

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