|
| Status |
Public on Mar 01, 2015 |
| Title |
Day4_4OHT_Rep1 |
| Sample type |
SRA |
| |
|
| Source name |
Differentiated embryonic stem cells
|
| Organism |
Mus musculus |
| Characteristics |
cell line: Brg1f/f; Actin-CreER
timepoint: Day 4
brg1 levels: Deleted
|
| Treatment protocol |
On Day2, cultures were treated with either 200 nM 4-OHT in tetrahydrofuran (THF) or THF alone. 4-OHT induces CreER mediated deleted of Brg1 floxed alleles.
|
| Growth protocol |
Mouse ES cells were cultured in feeder-free conditions under standard conditions. Mouse ES cells were aggregated into embryoid bodies (EB) and differentiated for two days in serum free media (Day2). Embryoid bodies were dissociated and reaggregated for 40 hours in the presence of VEGF, Activin A, and BMP4 to promote mesoderm differentiation (Day4).
|
| Extracted molecule |
polyA RNA |
| Extraction protocol |
RNA was extracted using TRIzol according to the manufacturer's instructions.
8 ug of total RNA was used as input material for the preparation of the RNA-Seq libraries, according to Illumina RNA Seq library kit with minor modifications. Briefly, mRNA was isolated using Dynabeads® mRNA Purification Kit (Invitrogen) followed by fragmentation (Ambion) and ethanol precipitation. First and second strand synthesis were performed followed by end repair, A-tailing, adapter ligation and size selection on a Beckman Coulter SPRI TE nucleic acid extractor. 200-400 bp dsDNA was enriched by 13 cycles of PCR with Phusion® High-Fidelity DNA Polymerase (NEB). Amplified libraries were sequenced on an Illumina HiSeq 2000 system.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 2000 |
| |
|
| Data processing |
Reads were aligned using Bowtie reporting uniquely aligned reads and allowing 2 bp mismatches
Generated .useq files using (USeq Version 8) Application Sam2USeq
USeq files were converted to BigWig format via GnomEx (https://hci-as3.hci.utah.edu/gnomex/gnomex.html)
Genome_build: mm9
Supplementary_files_format_and_content: Bigwig values are per base read depth scaled per million mapped reads.
|
| |
|
| Submission date |
Mar 23, 2013 |
| Last update date |
May 15, 2019 |
| Contact name |
Jeffrey Michael Alexander |
| E-mail(s) |
[email protected]
|
| Organization name |
Gladstone Institutes
|
| Lab |
Bruneau Lab
|
| Street address |
1650 Owens St
|
| City |
San Francisco |
| State/province |
CA |
| ZIP/Postal code |
94158 |
| Country |
USA |
| |
|
| Platform ID |
GPL13112 |
| Series (2) |
| GSE45446 |
Brg1 Modulates Enhancer Activation and Polycomb-mediated Repression in Mesoderm Differentiation [RNA-Seq] |
| GSE45448 |
Brg1 Modulates Enhancer Activation and Polycomb-mediated Repression in Mesoderm Differentiation |
|
| Relations |
| Reanalyzed by |
GSE80797 |
| SRA |
SRX254794 |
| BioSample |
SAMN01990871 |
