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Sample GSM2889293 Query DataSets for GSM2889293
Status Public on Dec 11, 2018
Title shNcor2 rep#1 (ATAC-seq)
Sample type SRA
Source name mouse embryonic stem cell
Organism Mus musculus
Characteristics cell type: mouse embryonic stem cell
cell line: E14
knockdown: shNcor2
Treatment protocol pLKO lentiviruses, generated with Plat-E cells, were added to ESCs and medium was changed after 8 hours. Cells were selected with 2 μg/ml puromycin for 2 days, and harvested after a further 2 days.
Growth protocol Mouse ESCs were grown in serum+LIF medium: DMEM-HIGH GLUCOSE (SH30022.01, HyClone), 15% fetal bovine serum (S1580-500, Biowest), 1% L-Glutamine (35050-061, GIBCO), 1% sodium pyruvate (25-000-CIR, Corning), 1% non-essential amino acids (25-025-CI, Corning), 0.5% penicillin-streptomycin solution (SV30010, HyClone), 0.1 mM 2-mercaptoethanol (21985-023, GIBCO), 0.01% mLIF (ESG1107, Millipore).
Extracted molecule genomic DNA
Extraction protocol ATAC-seq was performed as described (Buenrostro et al., 2015). In brief, a total of ~50,000 cells were washed once with 50 μl of cold PBS and resuspended in 50μl lysis buffer (10 mM Tris-HCl pH 7.4, 10 mM NaCl, 3 mM MgCl2, 0.2% (v/v) IGEPAL CA-630). The suspension of nuclei was then centrifuged for 10 min at 500g at 4°C, followed by the addition of 50 μl transposition reaction mix (25 μl TD buffer, 2.5 μl Tn5 transposase and 22.5 μl nuclease-free H2O) of Nextera DNA library Preparation Kit (96 samples) (FC-121-1031, Illumina). Samples were then PCR amplified and incubated at 37°C for 30min. DNA was isolated using a MinElute Kit (Qiagen). ATAC-seq libraries were first subjected to 5 cycles of pre-amplification. To determine the suitable number of cycles required for the second round of PCR the library was assessed by quantitative PCR as described (Buenrostro et al., 2015), and the library was then PCR amplified for the appropriate number of cycles. Libraries were purified with a Qiaquick PCR (Qiagen) column and were quantified using a KAPA Library Quantification kit (KK4824, Kapa Biosystems) according to the manufacturer’s instructions. Library integrity was checked by gel electrophoresis. Finally, the ATAC library was sequenced on a NextSeq 500 using a NextSeq 500 High Output Kit v2 (150 cycles) (FC-404-2002, Illumina) according to the manufacturer’s instructions.
According to the extract protocol
Library strategy ATAC-seq
Library source genomic
Library selection other
Instrument model Illumina NextSeq 500
Data processing bcl2fastq conversion software v2.17 used for basecalling.
Reads were aligned to the mouse (mm10) genome using bowtie2
Peaks were called using MACS2
Genome_build: mm10
Supplementary_files_format_and_content: Processed peak (BED) files and bigWig tracks
Submission date Dec 14, 2017
Last update date Dec 12, 2018
Contact name Jiangping He
Organization name Guangzhou Institutes of Biomedicine and Health (GIBH), CAS
Street address Kai yuan avenue 190
City GuangZhou
ZIP/Postal code 510530
Country China
Platform ID GPL19057
Series (2)
GSE108086 Epigenetic control of genes and transposable elements [ATAC-seq]
GSE108091 Epigenetic control of genes and transposable elements
BioSample SAMN08177864
SRA SRX3471729

Supplementary file Size Download File type/resource 158.7 Mb (ftp)(http) BW
GSM2889293_shNcor2.rep1.peaks.bed.gz 230.8 Kb (ftp)(http) BED
SRA Run SelectorHelp
Raw data are available in SRA
Processed data provided as supplementary file

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