|
| Status |
Public on Sep 06, 2016 |
| Title |
RNA siATF4 12h glucose C |
| Sample type |
SRA |
| |
|
| Source name |
β-cell_RNA siATF4 12h glucose
|
| Organism |
Rattus norvegicus |
| Characteristics |
cell line: INS-1E
cell type: beta-cell
|
| Treatment protocol |
One day prior to transfection INS-1E cells were seeded in antibiotic-free culture medium at a density of 800.000 cells/cm2. A transfection mix (750nM siRNA and 5% v/v DharmaFECT (Thermo) prepared in serum- and antibiotic-free OptiMEM media (Gibco) was added to the cells in a 1:5 ratio. The following day, the media was changed to standard RPMI, and the cells were incubated for 2 days and then switched to 5mM glucose medium for 24 h before exposure to (25 mM) or low (5mM) glucose for 12 h.
|
| Growth protocol |
INS-1E cells were cultured in RPMI 1640 w. 11 mM glucose supplemented with 5% Heat inactivated FCS, 50 uM β-MeOH, 1mM NaPyruvate, and 100 U/ml Penicilin + 100 mg/ml streptomycin. Cells were used between passages 60 and 90
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Following Isol-RNA lysis Reagent® (5-Prime) extraction and EconoSpin (Epoc Life) column purification of total RNA, polyadenylated RNA was isolated using poly-dT beads and RNA fragmentation and cDNA synthesis was performed according to the manufacturers (Truseq 2® , Illumina) instructions.
RNA-, DNase-, and ChIP-seq libraries were constructed using PentAdapters (Pentabase) essentially as previously described in (Nielsen R, Mandrup S, 2014, Methods in Enzymology 2014, Vol. 537, pp. 261-279).
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 1500 |
| |
|
| Description |
processed data file: Knockdowns_exon.txt
|
| Data processing |
alignment: RNA-seq reads were mapped to rn5 with STAR (Dobin, A., et al., Bioinformatics, 2013. 29(1): p. 15-21) using default parameters.
quantification: mRNA-seq coverage within exons of Ensembl (Rnor_5.0) genes were quantified using the iRNA-seq pipeline (Madsen et al, Nucl. Acids Res. 43 (6): e40), to asses mature transcript levels. EdgeR normalized readcounts are provided in Knockdowns_exon.txt
Genome_build: rn5
|
| |
|
| Submission date |
May 19, 2016 |
| Last update date |
May 15, 2019 |
| Contact name |
Susanne Mandrup |
| E-mail(s) |
s.mandrup@bmb.sdu.dk
|
| Phone |
+45 6550 2340
|
| Organization name |
University of Southern Denmark
|
| Department |
Biochemistry and Molecular Biology
|
| Street address |
Campusvej 55
|
| City |
Odense M |
| ZIP/Postal code |
5230 |
| Country |
Denmark |
| |
|
| Platform ID |
GPL18404 |
| Series (1) |
| GSE81628 |
Integrative genomics outlines a biphasic glucose response and a ChREBP-RORγ axis regulating proliferation in β-cells |
|
| Relations |
| BioSample |
SAMN05019589 |
| SRA |
SRX1774903 |
