|
| Status |
Public on Oct 26, 2021 |
| Title |
D4_BrmKO_high_BMP4_scRNA-seq |
| Sample type |
SRA |
| |
|
| Source name |
mouse ES cell derived cardiac mesoderm
|
| Organism |
Mus musculus |
| Characteristics |
cell type: mouse ES cell derived cardiac mesoderm
genotype: Brm-/-
timepoint: Day 4
|
| Treatment protocol |
mouse ESC lines were differentiated to cardiomyocytes using the Kattman et al 2011 Cardiomyocyte Differentiation protocol.
|
| Growth protocol |
mouse ESCs were maintained in DMEM media supplemented with LIF and serum
|
| Extracted molecule |
polyA RNA |
| Extraction protocol |
Cells on day 4, day 6 and day 10 were singularized using Trypsin 0.25% and 10,000 cells were loaded on a 10X Chromium chip
Chromium Single Cell 3′ GEM, Library & Gel Bead Kit v2 was used to construct libraries
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina NovaSeq 6000 |
| |
|
| Data processing |
Libraries were sequenced on Illumina Novaseq
Raw sequencing reads were converted to FASTQ files using Cellranger 2.0.2 mkfastq
Reads were aligned to mm9 using the Cellranger 2.0.2 count
Cellranger 2.0.2 aggr was used to aggregate libraries from WT and BRM KO cells at normal BMP4 concentration or combined with high BMP4 concentrations
Aggregated gene expression matrix was processed in Seurat 2.3.4. Briefly, cells with UMI counts between 10,000 and 80,000 were retained. Gene expression was normalized, scaled and mitochondrial reads and cell cycle genes were regressed. Cells were clustered based on the top 25-30 prinicipal components.
URD (v1.0.2) was used for trajectory analysis. Seurat matrix was converted to an URD object, cell-to-cell transition probabilities were calculated by setting number of nearest neighbors to 211 and sigma of 8.
Pseudotime was then calculated by running 80 flood simulations with POU5F1+ clusters as the ‘root’ cells. Next, all day 10 clusters were set as ‘tip’ cells and biased random walks were simulated from each tip to build an URD tree.
Genome_build: mm9
Supplementary_files_format_and_content: Processed Seurat expression matrix as a csv file, with columns as cell barcodes and rows as genes
|
| |
|
| Submission date |
May 09, 2020 |
| Last update date |
Oct 26, 2021 |
| Contact name |
Benoit Bruneau |
| E-mail(s) |
[email protected]
|
| Organization name |
Gladstone Institute of Cardiovascular Disease
|
| Street address |
1650 Owens Street
|
| City |
San Francisco |
| State/province |
CA |
| ZIP/Postal code |
94158 |
| Country |
USA |
| |
|
| Platform ID |
GPL24247 |
| Series (2) |
| GSE150185 |
Day4_Day6_Day10_normalBMP4_highBMP4_single_cell_RNAseq |
| GSE150186 |
Brahma safeguards canalization of cardiac mesoderm differentiation |
|
| Relations |
| BioSample |
SAMN14860211 |
| SRA |
SRX8297783 |
