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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
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)
GSE79738 Requirement for flap endonuclease 1 (FEN1) to maintain genomic stability and transcriptional gene silencing in Arabidopsis
Relations
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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