|
| Status |
Public on Jan 25, 2019 |
| Title |
NS_C |
| Sample type |
SRA |
| |
|
| Source name |
Bronchial epithelial cells
|
| Organism |
Homo sapiens |
| Characteristics |
cell type: Bronchial epithelial cells
stimulation: Non-stimulated control
|
| Treatment protocol |
Once fully differentiated, 1μM JAK1 inhibitor in DMSO or DMSO were added apically for 30 minutes in 200μL medium that was removed before stimulation. Cells were stimulated for 24h by basolateral addition of human recombinant proteins IL-6 (150 ng/ml; R&D Systems) and sIL-6R (150 ng/ml; PeproTech) diluted in BEBM.
|
| Growth protocol |
Primary human bronchial epithelial cells (HBECs; Lonza) were expanded to passage 2 in BEGM Bronchial Epithelial Cell Growth Medium (Lonza). These cells were then seeded on 0.4 μm Corning® HTS Transwell®-24 well permeable supports (Sigma-Aldrich) coated with PureCol (Advanced BioMatrix) and differentiated at Air-Liquid Interface (ALI) using BEBM (Lonza) according to manufacturer’s protocol.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Cells were lysed in QIAzol Lysis Reagent (Qiagen), scraped off the wells and briefly vortexed. RNA was isolated using the RNeasy Mini Kit (Qiagen) following the manufacturer's instruction and RNA integrity was assessed (Bioananalyzer 2100, Agilent).
RNA was diluted to 10 ng/μl and converted to mRNA libraries using the TruSeq Stranded mRNA kit (Illumina) with dual indexing following standard instructions but with 5 min fragmentation time. Libraries were validated on the Fragment Analyzer platform (AATI) using a standard sensitivity NGS fragment analysis kit, and concentrations were determined using the Quant-iT dsDNA High Sensitivity assay kit on the Qubit fluorometer (Thermo Fisher Scientific). Sample libraries were pooled in equimolar concentrations, diluted and denatured according to Illumina guidelines.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina NextSeq 500 |
| |
|
| Description |
Sample 3 (Study 1 - IL-6 vs T2 study)
Study1_combined.counts
Study1_combined.gene.sf.tpm
|
| Data processing |
The two studies' fastq files were processed separately using bcbio-nextgen (version 0.9.9a-1ccf093 (study 1) / 0.9.7 (study2)) where reads were mapped to the human genome build hg38 (GRCh38.79) using hisat2 (version 2.0.4 (study 1) / 2.0.2-beta (study 2)).
Gene level quantifications, counts and transcript per million (TPM), were generated with featurecounts (version 1.4.4 (study 1) / 1.4.4 (study 2)) and sailfish (version 0.10.1 (study 1) / 0.9.0 (study 2)), respectively, all within bcbio.
Genome_build: hg38
Supplementary_files_format_and_content: Counts-files contains gene level quantifications from featurecounst used for DE analysis. TPM-files contains gene level quantifications from sailfish.
|
| |
|
| Submission date |
Apr 16, 2018 |
| Last update date |
Jan 25, 2019 |
| Contact name |
Elisabeth Israelsson |
| E-mail(s) |
[email protected]
|
| Organization name |
AstraZeneca, IMED RIA
|
| Department |
Translational Biology
|
| Street address |
Pepparedsleden 1
|
| City |
Molndal |
| ZIP/Postal code |
SE-431 83 |
| Country |
Sweden |
| |
|
| Platform ID |
GPL18573 |
| Series (1) |
| GSE113185 |
IL-6 trans-signaling induced gene expression in airway epithelial cells |
|
| Relations |
| BioSample |
SAMN08937538 |
| SRA |
SRX3942531 |
