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GEO help: Mouse over screen elements for information. |
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| Status |
Public on Jan 25, 2019 |
| Title |
ATAC WT NIH-3T3 |
| Sample type |
SRA |
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| Source name |
NIH-3T3
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| Organism |
Mus musculus |
| Characteristics |
cell line: NIH-3T3
cell type: mouse fibroblasts
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| Treatment protocol |
50'000 cells were collected.
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| Growth protocol |
NIH-3T3 cells were routinely cultured at 37°C and 5% CO2 in DMEM, supplemented with 10% fetal bovine serum and 1% penicillin/streptomycin. Cells were grown in 100 mm cell culture dishes up to a confluence of 90% and split 1:6 every 3-4 days.
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
Cells were pelleted and washed with 1X ice cold PBS at 800g for 5 min. Cells were gently resuspended in 50 μl of ice-cold ATAC lysis buffer (10 mM Tris-HCl pH 7.4, 10 mM NaCl, 3 mM MgCl2, 0.1% NP40), and immediately pelleted at 800g for 10 min at 4°C. To transpose open chromatin regions, cells were resuspended in 50 μl of transposition reaction mix containing 0.5 μM of Tn5 transposase (in-house prepared) in TAPS-DMF buffer (10 mM TAPS-NaOH, 5 mM Mgcl2, 10% DMF) and incubated at 37°C for 30 min. The transposed DNA was purified using a DNA purification kit and eluted in 12 μl of water.
A 65 μl PCR reaction was setup with 10 μl of transposed DNA, 0.5 μM of forward primer Ad1_noMX, 0.5 μM of multiplexing reverse primer Ad2.x (Buenrostro et al. 2013), 0.6x SYBR Green I, and 1x PCR Master Mix. The samples were thermocycled at 72°C for 5 minutes, 98°C for 30 s, followed by 12 cycles of 98°C for 10 s, 63°C for 30 s and 72°C for 1 min. The amplified ATAC libraries were purified using a DNA purification kit and size selected using Agencourt AMPure beads (0.55X unbound fraction followed by 1.2X bound fraction).
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| Library strategy |
ATAC-seq |
| Library source |
genomic |
| Library selection |
other |
| Instrument model |
Illumina NextSeq 500 |
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| Data processing |
Reads were aligned to the mouse reference genome mm10 using STAR 2.5.3a with settings ‘-- alignMatesGapMax 2000 --alignIntronMax 1 --alignEndsType EndtoEnd -- outFilterMultimapNmax 1’
Duplicate reads were removed with Picard and reads not mapping to chromosomes 1-19, X, or Y were removed
Peaks were called with MACS 2.1.1.20160309 with settings ‘-f BAMPE -g mm’.
Genome_build: mm10
Supplementary_files_format_and_content: bigWig files were generated by merging replicate bam files with SAMtools followed by the deepTools functions bamCoverage (with setting --normalizeUsingRPKM). log2-ratio bigWig files were generated using the deepTools function bamCompare based on merged replicates for each TF overexpression sample over merged replicates for control (rtTA3G).
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| Submission date |
Sep 11, 2018 |
| Last update date |
Jan 27, 2019 |
| Contact name |
David Michael Suter |
| Organization name |
École polytechnique fédérale de Lausanne
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| Department |
Institute of Bioengineering
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| Lab |
UPSUTER
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| Street address |
EPFL SV-IN Station 19
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| City |
Lausanne |
| ZIP/Postal code |
1015 |
| Country |
Switzerland |
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| Platform ID |
GPL19057 |
| Series (2) |
| GSE119781 |
Mitotic chromosome binding predicts transcription factor properties in interphase [ATAC-Seq] |
| GSE119784 |
Mitotic chromosome binding predicts transcription factor properties in interphase |
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| Relations |
| BioSample |
SAMN10032313 |
| SRA |
SRX4669275 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM3383675_ATAC_3T3_WT.bw |
181.3 Mb |
(ftp)(http) |
BW |
| GSM3383675_ATAC_3T3_WT_BAMPE_peaks.narrowPeak.gz |
1.6 Mb |
(ftp)(http) |
NARROWPEAK |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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