|
| Status |
Public on Sep 04, 2015 |
| Title |
ES-CTR [RNA-Seq] |
| Sample type |
SRA |
| |
|
| Source name |
Undifferentiated ES cells
|
| Organism |
Mus musculus |
| Characteristics |
cell line: E14Tg2A
cell type: Embryonic stem cells (ESCs)
genotype/variation: Control
|
| Treatment protocol |
In our hands, generation of highly‐enriched population of MES cells was strongly influenced both by the dose and the batch of BMP4. We performed full cardiomyocyte differentiation experiments using 32 different amounts of BMP4 from 0.1 to 0.8 ng/ml with every purchased BMP4. Typically, we obtained the best differentiation results with 0.1–0.3 ng/ml of BMP4. We also noticed that the amount of MES cells seeded to obtain cardiac precursor cells (CP) was crucial to obtain a high percentage of beating cardiomyocytes, with the best results obtained when 125,000–150,000 MES cells were seeded in a 96-well plate.
|
| Growth protocol |
E14Tg2A cells were cultured as previously described (Morey et al., 2012) in 2i media supplemented with LIF produced in house) as previously described (Ying et al., 2003) for at least two weeks (8 passages). We took advantage of a recently reported protocol for differentiating ESC into a highly homogenous population of beating cardiomyocytes (CM) (Kattman et al., 2011). Interestingly, this protocol also permits early steps of cardiac differentiation, like formation of early cardiac-mesoderm precursors cells (MES) and cardiac precursor (CP) cells, to be studied.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Chromatin from ESC and MES was immunoprecipitated with 5 ug of antibody as described in Morey et al 2012. RNA was extracted with the RNA extraction kit (QIAGEN).
Libraries were prepared according to Illumina instructions.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 2000 |
| |
|
| Description |
(RNAseq) Control
|
| Data processing |
ChIPseq analysis: Alignment: Sequence reads were mapped to the Mouse genome (mm9, July 2007) using BOWTIE (PubMed ID 19261174) with the option -m 1. RNAseq analysis: Alignment: The reads were mapped against the mm9 mouse genome assembly using TopHat (PubMed ID 22383036) with the option ‐g 1.
Peak-calling: Peak detection was performed with MACS (PubMed ID: 18798982). ChIP profiles were produced by MACS in BedGraph. Peaks were reported in BED format.
Genome_build: mm9
Supplementary_files_format_and_content: BedGraph files report genome-wide ChIP or RNAseq profiles. BED files report signal enriched regions.
|
| |
|
| Submission date |
Apr 14, 2015 |
| Last update date |
May 15, 2019 |
| Contact name |
Enrique Blanco |
| E-mail(s) |
enrique.blanco@crg.eu
|
| Phone |
+34 93 316 01 00
|
| Organization name |
Center for Genomic Regulation (CRG)
|
| Department |
Gene Regulation, Stem Cells and Cancer
|
| Lab |
Epigenetic Events in Cancer (L. Di Croce's lab)
|
| Street address |
Dr. Aiguader 88
|
| City |
Barcelona |
| ZIP/Postal code |
08003 |
| Country |
Spain |
| |
|
| Platform ID |
GPL13112 |
| Series (1) |
| GSE67868 |
The Polycomb protein Pcgf2/Mel18 regulates mesoderm cell fate-specification of embryonic stem cells through multiple mechanisms |
|
| Relations |
| BioSample |
SAMN03482386 |
| SRA |
SRX994832 |
