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Sample GSM2782415 Query DataSets for GSM2782415
Status Public on Jul 15, 2018
Title ATAC-seq from Gatad2a system, Day4
Sample type SRA
 
Source name Gatad2a cell line, Day4 post DOX induction
Organism Mus musculus
Characteristics tissue: Gatad2a-/- cell line
stage of reprogramming: Day4
genotype: Gatad2a-/-
Treatment protocol Reprogramming dynamics samples were generated by adding Dox to Secondary MEF. ES and iPSC samples were harvested after 3-4 days of growth in N2B27 2i\LIF medium. Reprogramming samples of days 3-8, ES and iPSC samples were plated on irradiated human foreskin fibroblasts (HFF).
Growth protocol Maintenance and reprogramming (Day 3 and beyond) of murine naïve pluripotent cells were conducted in bovine serum-free N2B27-based media or KSR media: N2B27- based media: 500ml KO-DMEM (Invitrogen), 5ml N2 supplement (Invitrogen; 17502048), 5ml B27 supplement (Invitrogen; 17504044), 15% knockout serum replacement (Invitrogen – 10828), 1mM glutamine (Invitrogen), 1% nonessential amino acids (Invitrogen), 0.1mM β-mercaptoethanol (Sigma), penicillin-streptomycin (Invitrogen), 5mg/ml BSA (Sigma). KSR media: 500 ml DMEM (Invitrogen) , 1mM glutamine (Invitrogen), 1% nonessential amino acids (Invitrogen), 0.1mM β-mercaptoethanol (Sigma), penicillin- streptomycin (Invitrogen). Naïve conditions for murine PSCs included 10μg recombinant human LIF (Millipore; LIF1005). Where indicated 2i was added 48 hours after OSKM induction: small-molecule inhibitors CHIR99021 (CH, 3μM- Axon Medchem) and PD0325901 (PD 0.2 or 1μM - Axon Medchem).
Extracted molecule genomic DNA
Extraction protocol 50,000 cells were centrifuged (500g, 3 minutes), then washed using 50 μL of cold PBS, and centrifuged again (500g for 3 min). Cells were lysed using cold lysis buffer (10 mM Tris-HCl, pH 7.4, 10 mM NaCl, 3 mM MgCl2 and 0.1% IGEPAL CA-630). Immediately after lysis, nuclei were spun at 500g for 10 min using a refrigerated centrifuge. Next, the pellet was re-suspended in the transposase reaction mix (25 μL 2× TD buffer, 2.5 μL transposase (Illumina) and 22.5 μL nuclease-free water). The transposition reaction was carried out for 30 minutes at 37 °C and immediately transfered to ice. The sample was then purified using Qiagen MinElute kit.
Following purification, the library fragments were amplified using custom Nextera PCR primers 1 and 2 for a total of 12 cycles. Following PCR amplification the libraries were purified using a Qiagen MinElute Kit.
 
Library strategy ATAC-seq
Library source genomic
Library selection other
Instrument model Illumina NextSeq 500
 
Data processing We used Illumina CASAVA 1.8.2 software to generate fastq files.
Reads were aligned to mm10 mouse genome using Bowtie2 with the parameter -X2000 (allowing fragments up to 2 kb to align).
Duplicated aligned reads were removed using Picard MarkDuplicates tool with the command REMOVE_DUPLICATES=true.
To identify chromatin accessibility signal we considered only short reads (≤ 100bp) that correspond to nucleosome free region.
To detect and separate accessible loci in each sample, we used MACS version 1.4.2-1 with --call-subpeaks flag (PeakSplitter version 1.0).
93,137 enhancers with co-localized ATAC-seq and H3K27ac peaks were identified.
For each promoter and each enhancer we report here the read depth in a 50-bp bins, 1 Kb around the TSS or 300 bp around the enhancer summit.
Genome_build: mm10
Supplementary_files_format_and_content: Promoter "matrix" files contain for each TSS, the number of reads in 50bp bins, spanning 1Kb window around the TSS. 1st column - gene esemble ID, columns 2-21 - read num in 50bp bins. Enhancer "matrix" files contain for each identified enhancer, the number of reads in 50bp bins, spanning 300bp window around the enhancer summit. 1st column - enhaner genomic interval. 2nd column - enhancer ID number, columns 3-8 - read num in 50bp bins.
 
Submission date Sep 13, 2017
Last update date May 15, 2019
Contact name Noa Novershtern
E-mail(s) noa.novershtern@weizmann.ac.il
Organization name Weizmann Institute of Science
Department Molecular Genetics
Street address Weizmann Institute
City Rehovot
ZIP/Postal code 7610001
Country Israel
 
Platform ID GPL19057
Series (2)
GSE102518 High-Resolution Dissection of Conducive Somatic Cell Reprogramming to Naïve Ground State Pluripotency in Mbd3/NuRD and Gatad2a/NuRD Depleted Systems
GSE103821 High-Resolution Dissection of Conducive Somatic Cell Reprogramming to Naïve Ground State Pluripotency in Mbd3/NuRD and Gatad2a/NuRD Depleted Systems [ATAC-Seq]
Relations
BioSample SAMN07638588
SRA SRX3182578

Supplementary file Size Download File type/resource
GSM2782415_Gatad2a_ATAC_Day4.enhancer_150.matrix.txt.gz 1.8 Mb (ftp)(http) TXT
GSM2782415_Gatad2a_ATAC_Day4.prom_500.matrix.txt.gz 2.3 Mb (ftp)(http) TXT
SRA Run SelectorHelp
Raw data are available in SRA
Processed data provided as supplementary file

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