|
| Status |
Public on Jan 27, 2022 |
| Title |
d0d4_earlydiff |
| Sample type |
SRA |
| |
|
| Source name |
differentiating embryonic stem cells
|
| Organism |
Mus musculus |
| Characteristics |
time: day 0 through day 4
developmental stage: embryonic stem cells undergoing differentiation to early gastrulation
|
| Treatment protocol |
mESCs were removed from feeder cells and cultured in suspension in serum-free differentiation media (SFD) from day0 to day 2. At day 2, four different primitive streak induction treatments were established by adding growth factors Wnt3a (25 ng/mL) and Activin A (9 ng/mL) alone or with one of the following additional growth factors: Noggin (150 ng/mL) Gsk3 inhibitor (10 mM), or Bmp4 (0.5 ng/mL).
|
| Growth protocol |
GFP-Bracyury mESC cells were maintained on mouse embryonic feeder (mEF) cells in Dulbecco's Modified Eagles' Medium (DMEM; Gibco) containing 15% FBS (Sigmal-Aldrich), 1% PSG (Gibco), LIF (MTI Global Stem; 1,000 U/mL), and 2-mercaptoethanol (Sigma; 0.1mM).
|
| Extracted molecule |
polyA RNA |
| Extraction protocol |
Single-cell samples were prepared by dissocaiting cells in TrypLE before strained through a 40 uM cell strainer.
Sample barcoding and library preparation was carried out using the MULTI-seq protocol (McGinnis et al. 2019).
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina NovaSeq 6000 |
| |
|
| Data processing |
Sequencing alignment to the mm10 reference genome was performed using CellRanger (version 3.1), and spliced, unspliced, and ambiguous read counts were extracted using Velocyto.
Sample barcode classification was performed using the deMULTIplex R package, and doublets and negatives were removed.
Normalization, log transformation, and other data processing carried out in Scanpy (version 1.4.5; Wolf et al., 2018).
Genome_build: mm10
Supplementary_files_format_and_content: Processed data, including cluster assignments, provided in h5ad format. Cell metadata (names, timepoint, treatment, cluster, etc.) are stored in .obs. Raw counts data is stored in .raw. Normalized and log-transformed data is stored in .X.
|
| |
|
| Submission date |
Jun 11, 2021 |
| Last update date |
Jan 27, 2022 |
| Contact name |
Patrick Cahan |
| E-mail(s) |
patrick.cahan@jhmi.edu
|
| Organization name |
Johns Hopkins University School of Medicine
|
| Department |
ICE and Biomedical Engineering
|
| Lab |
Cahan Lab
|
| Street address |
733 North Broadway, MRB 653
|
| City |
Baltimore |
| State/province |
MD |
| ZIP/Postal code |
21205 |
| Country |
USA |
| |
|
| Platform ID |
GPL24247 |
| Series (1) |
| GSE177051 |
Reconstruction of dynamic regulatory networks reveals signaling-induced topology changes associated with germ layer specification |
|
| Relations |
| BioSample |
SAMN19671526 |
| SRA |
SRX11122147 |
