|
| Status |
Public on Jul 15, 2024 |
| Title |
A6_Ezh2I_3_S29 |
| Sample type |
SRA |
| |
|
| Source name |
mouse embryonic fibroblasts
|
| Organism |
Mus musculus |
| Characteristics |
cell type: mouse embryonic fibroblasts
genotype: C57/B16 background
treatment: Ezh2i
time: Day 4
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
We transduced mouse embryonic fibroblasts with a CRISPRa-VPR–mCherry construct and assessed more immediate and longer term effects of Fra2 over-expression by performing OMNI-ATACseq on FACS purifed mCherry+/THY1+ cells 4 and 11 days following lentiviral infection with Fra2 gRNAs or scrambled control.For Ezh2 inbition:Following three days of treatment with the Ezh2 inhibitor (Sigma-Aldrich, GSK343) at 4μM and a washout day, MEFs were submitted to the ATAC-seq assay.
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| |
|
| Library strategy |
OTHER |
| Library source |
genomic |
| Library selection |
other |
| Instrument model |
NextSeq 2000 |
| |
|
| Description |
ATAC_Fosl2_Ezh2i_peaks.gtf
ATAC_Fosl2_Ezh2i.counts.txt
|
| Data processing |
ATAC-seq: Peak calling was performed using the MACS2 callpeak command with the following parameters “-g mm –shift -100 –extsize 200 –nomodel –call-summits –nolambda –keep-dup all -p 0.01”. Then, peak summits were extended by 250 bp on both sides to a final width of 501 bp. Peaks were filtered for mm10 blacklisted regions (https://www.encodeproject.org/annotations/ENCSR636HFF/).
As in Corces et al. 2020 per sample overlapping peaks were handled using an iterative removal approach. That is in case of two overlapping peaks the most significant peak is kept and the other removed. This process is performed in an iterative manner, so all peaks have been kept or removed based on their overlap and significance score. This method results in a set of fixed-width peaks per sample.
Next, significance peak scores “(-log10(p-value))” were converted to a score per million value by dividing each individual peak score by the sum of all of the peak scores in the given sample divided by 1 million. This normalized score per million value corrects the original peak calling scores for sample sequencing depth and quality, as higher quality samples yield higher number of peaks and higher significance scores overall. Thus, scores per million allow the direct comparison of peaks across biological replicates.
Next, we compiled a condition - MEF, mESC- and sequencer specific - BGI, Illumina - peak set containing reproducible peaks observed in a particular cell type and sequencing platform. For that, we generated a cumulative peak set per cell type and sequencing platform and performed the same iterative approach as mentioned above. Therefore, maintaining the peak with the highest significance or score per million. Only peaks with a score per million value >=3 observed in at least two samples (minimal overlap 50%) were further considered.
Assembly: mm10
Supplementary files format and content: Matrix table with counts per peaks in (.txt) format, samples mapping in bigwig format (.bw) and peaks called per individual sample as (.txt) format.
Library strategy: Omni ATAC-seq
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| |
|
| Submission date |
Jun 11, 2024 |
| Last update date |
Jul 15, 2024 |
| Contact name |
Christian Nefzger |
| Organization name |
The University of Queensland
|
| Department |
Institute for Molecular Biology
|
| Lab |
Cellular Reprogramming and Ageing
|
| Street address |
306 Carmody Road
|
| City |
Brisbane |
| State/province |
Queensland |
| ZIP/Postal code |
4067 |
| Country |
Australia |
| |
|
| Platform ID |
GPL30172 |
| Series (1) |
| GSE217780 |
Fosl2 overexpression and Ezh2 inhibition in mouse embryonic fibroblasts |
|
| Relations |
| BioSample |
SAMN41786539 |
| SRA |
SRX25044394 |
