|
| Status |
Public on Oct 09, 2015 |
| Title |
d0_pellet4_rnaseq |
| Sample type |
SRA |
| |
|
| Source name |
C32 iPSC_d0_pellet_rnaseq
|
| Organism |
Homo sapiens |
| Characteristics |
cell line: C32 iPSC
days cultured: 0
cell type: mini-kidney derived from human induced pluripotent stem cells (C32 iPSC)
|
| Treatment protocol |
When cells reached to 40-50 % of confluent, cells were treated with 8 mM of CHIR99021 in APEL basal medium (STEMCELL Technologies) supplemented with Antibiotic-Antimycotic (Life Technologies) for 4 days, followed by FGF9 (200 ng mL-1) and Heparin (1 mg mL-1) for another 3 days, with changing medium every second day. Then, cells were collected and dissociated into single cells using Trypsin or TrypLE select (Life Technologies). Cells (0.5-1 × 10^6) were spun down at ×400g for 2 min to form a pellet and then placed onto a Transwell 0.4 mm pore polyester membrane (#CLS3450 Corning). A pellet was treated with 5 mM of CHIR99021 in APEL for 1 h, and then cultured with FGF9 (200 ng mL-1) and Heparin (1 mg mL-1) for 5 days, followed by another 6-13 days in APEL basal medium, with changing medium every second day.
|
| Growth protocol |
Human iPSCs were plated on a Matrigel-coated (Millipore) culture dish and cultured in MEF-conditioned hES medium. Then, cells were again plated on a Matrigel-coated at 5,000 cells/cm2.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted from cells using Purelink RNA mini kit (Life Technologies)
TruSeq stranded total RNA Ribo Zero Gold library prep (Illumina); RNA-seq; 1x76bp
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina NextSeq 500 |
| |
|
| Data processing |
Basecalling: Illumina Real-Time Analysis (RTA) v2.1.3
bcl to fastq conversion, including trimmming of adapters: Illumina bcl2fastq2 v2.1.4
Mapping: STAR v2.3.0e
Read counts per gene: htseq-count (HTSeq-0.6.1), with Illumina iGenomes UCSC hg19 annotation
Normalised counts: DESeq2
Genome_build: hg19
Supplementary_files_format_and_content: comma-separated values of raw counts and normalised counts (all replicates)
|
| |
|
| Submission date |
Jun 22, 2015 |
| Last update date |
May 15, 2019 |
| Contact name |
Melissa H Little |
| E-mail(s) |
[email protected]
|
| Organization name |
Murdoch Childrens Research Institute
|
| Department |
Cell Biology
|
| Lab |
Kidney Development, Disease and Regeneration
|
| Street address |
Flemington Road
|
| City |
Parkville |
| State/province |
Victoria |
| ZIP/Postal code |
3052 |
| Country |
Australia |
| |
|
| Platform ID |
GPL18573 |
| Series (1) |
| GSE70101 |
Mini-kidneys from human pluripotent cells model normal human fetal kidney development and response to nephrotoxicity |
|
| Relations |
| SRA |
SRX1059389 |
| BioSample |
SAMN03774849 |
