 |
 |
GEO help: Mouse over screen elements for information. |
|
| Status |
Public on Jun 30, 2020 |
| Title |
Thymus_Fibroblasts_SteadyState_ATAC_1 |
| Sample type |
SRA |
| |
|
| Source name |
Murine structural cells
|
| Organism |
Mus musculus |
| Characteristics |
organ: Thymus
cell type: GP38posCD31neg
condition: steady_state
age: 8 to 12 weeks
strain: C57BL/6J
|
| Treatment protocol |
not applicable
|
| Growth protocol |
C57BL/6J mice, bred and maintained under specific pathogen free conditions at the Institute of Molecular Biotechnology (IMBA) of the Austrian Academy of Sciences in Vienna, Austria. 8 to 12 week old mice (males only ) were used for the steady state characterization
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Organs were digested with Accumax and structural cells isolated using FACS
Chromatin accessibility mapping by ATAC-seq was performed as previously described (Buenrostro et al., 2013; Corces et al., 2016), with minor adaptations. In each experiment, a maximum of 50,000 sort-purified cells were collected at 300 g for 5 min at 4 °C. After centrifugation, the pellet was carefully resuspended in the transposase reaction mix (12.5 µl 2xTD buffer, 2 µl TDE1 (Illumina), 10.25 µl nuclease-free water, and 0.25 µl 1% digitonin (Promega)) for 30 min at 37 °C. Following DNA purification with the MinElute kit eluting in 11 µl, 1 µl of the eluted DNA was used in a quantitative PCR reaction to estimate the optimum number of amplification cycles. The remaining 10 μl of each library were amplified for the number of cycles corre-sponding to the Cq value (i.e., the cycle number at which fluorescence has increased above background levels) from the qPCR (rounded up). Library amplification was followed by SPRI (Beckman Coulter) size selection to exclude fragments larger than 1,200 bp. DNA concentration was measured with a Qubit fluo-rometer (Life Technologies). Library amplification was performed using custom Nextera primers (Buenrostro et al., 2013). The libraries were sequenced by the Biomedical Sequencing Facility at CeMM using the Illumina HiSeq 3000/4000 platform and the 50-bp single-end configuration.
|
| |
|
| Library strategy |
ATAC-seq |
| Library source |
genomic |
| Library selection |
other |
| Instrument model |
Illumina HiSeq 4000 |
| |
|
| Description |
open chromatin (ATAC-seq)
ATAC-seq sample of Thymus GP38posCD31neg cells
|
| Data processing |
Adapter trimming with trimmomatic (version 0.32) with settings LEADING:3 TRAILING:3 SLIDINGWINDOW:4:10 MINLEN:36
Genome alignment using bowtie (2.2.4) with settings --very-sensitive
Filtered reads to primary alignments with MAPQ > 30 using sambamba (0.7.0)
Called peaks using MACS2 (2.7.6)
Counted reads in consensus peak set over all samples using GenomicAlignments in R (3.2.3)
Normalized to counts per million (CPM) and transformed to log2(CPM) using Limma Voom in R
Regressed out immune cell signatures using Limma removeBatchEffect in R
Genome_build: mm10
Supplementary_files_format_and_content: Matrix of read counts (CSV), read counts of each peak in the consensus peak set for each sample
Supplementary_files_format_and_content: Matrix of log2(CPM) values (CSV), log2(CPM) after regressing out immune signatures of each peak in the consensus peak set for each sample
|
| |
|
| Submission date |
Mar 06, 2020 |
| Last update date |
Jun 30, 2020 |
| Contact name |
Christoph Bock |
| E-mail(s) |
cbock@cemm.oeaw.ac.at
|
| Organization name |
CeMM Research Center for Molecular Medicine of the Austrian Academy of Sciences
|
| Street address |
Lazarettgasse 14
|
| City |
Vienna |
| ZIP/Postal code |
1090 |
| Country |
Austria |
| |
|
| Platform ID |
GPL21103 |
| Series (2) |
| GSE134648 |
Structural cells are key regulators of organ-specific immune response |
| GSE134663 |
Structural cells are key regulators of organ-specific immune response |
|
| Relations |
| BioSample |
SAMN14322442 |
| SRA |
SRX7866261 |
| Supplementary data files not provided |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data are available on Series record |
|
|
|
|
 |
