|
| Status |
Public on Jun 15, 2016 |
| Title |
Kit_1.5h_CaptureC_merge |
| Sample type |
SRA |
| |
|
| Source name |
Kit_1.5h
|
| Organism |
Mus musculus |
| Characteristics |
cell line: G1E ER4
treatment: estradiol 13h
transgene: stably expressing YFP-MD
nocodazole: 1.5h release
promoter anchor: Kit promoter
raw file ids merged: 825_979_973
|
| Treatment protocol |
Except where indicated in the text as uninduced, we induced cells to mature with 100nM estradiol to activate GATA1-ER. During estradiol induction, we simultaneously treated cells with nocodazole (200ng/ml) for 7h-13h, washed once, and replated into fresh medium lacking nocodazole for varying times (40min-360min), ensuring all samples are exposed to estradiol for the same duration of 13h. We fixed cells with 1% formaldehyde, stained with DAPI, and sorted on a BD FACSAria based on YFP-MD and DAPI signal.
|
| Growth protocol |
G1E erythroblasts were previously derived through deletion of GATA1 in mouse embryonic stem cells, followed by in vitro differentiation (Weiss, Yu, and Orkin 1997). We cultured a sub-line of G1E cells, G1E-ER4, in which GATA1-ER was retrovirally transduced (referred to in main text as “G1E GATA1-ER"), as previously described (Weiss, Yu, and Orkin 1997). We retrovirally transduced G1E-ER4 cells with the YFP-MD construct (Kadauke et al. 2012) and sorted for a pool of YFP-positive cells.
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
After cell fixation with 1% formaldehyde for 10min and cell sorting as described above, Capture-C was performed with a double-capture procedure (Davies et al. 2016). Chromatin was digested using DpnII. We used biotin labelled DNA oligos to pull down the target restriction fragments.
Capture-C libraries were sequenced on Nextseq500 with 2x75 bp paired end sequencing.
|
| |
|
| Library strategy |
OTHER |
| Library source |
genomic |
| Library selection |
other |
| Instrument model |
Illumina NextSeq 500 |
| |
|
| Description |
Kit promoter anchor
825_979_973_Kit
|
| Data processing |
Library strategy: Capture-C
The raw reads were processed using published scripts (https://github.com/telenius/captureC/releases).
Custom scripts in R to normalize data by the total number of reads representing fragments ligated to the anchor region in the library.
For each time point and anchor, used custom scripts in R to combine reads representing fragments ligated to the anchor region in the library into one .bedgraph file merged across restriction digestion and/or capture replicates.
Genome_build: mm9
Supplementary_files_format_and_content: .bedgraph files containing coverage of reads (read counts of ligation products to given anchor region, per thousand total reads that represent ligation products to given anchor region).
|
| |
|
| Submission date |
Jun 13, 2016 |
| Last update date |
May 15, 2019 |
| Contact name |
Ross Hardison |
| E-mail(s) |
rch8@psu.edu
|
| Organization name |
Pennsylvania State University
|
| Street address |
303 Wartik Lab
|
| City |
University Park |
| State/province |
PA |
| ZIP/Postal code |
16802 |
| Country |
USA |
| |
|
| Platform ID |
GPL19057 |
| Series (2) |
| GSE83290 |
A hyperactive transcriptional state marks genome reactivation at the mitosis-G1 transition [CaptureC] |
| GSE83293 |
A hyperactive transcriptional state marks genome reactivation at the mitosis-G1 transition |
|
| Relations |
| BioSample |
SAMN05234762 |
| SRA |
SRX1839068 |
