|
Status |
Public on May 27, 2014 |
Title |
S1_RNA-Seq |
Sample type |
SRA |
|
|
Source name |
MCF7_Nutlin stimulated
|
Organism |
Homo sapiens |
Characteristics |
cell line: MCF7 cell type: breast cancer cells stimulated with: 5μM Nutlin-3a for 24hr
|
Treatment protocol |
The next day, cells were either stimulated with 5μM Nutlin-3a (in case of p53-WT cells) or left untreated.
|
Growth protocol |
Wild-Type (WT) MCF-7 cells were plated onto 24-well plates (60000 cells/well) in RPMI medium (+ L-glutamaat, Gibco) supplemented with 10% fetal bovine serum, 0.4mM sodium pyruvate (Gibco), 100μm/ml penicillin/streptomycin (Invitrogen), 1x non-essential aminoacids (Gibco) and 10μg/ml Insulin (Sigma).
|
Extracted molecule |
total RNA |
Extraction protocol |
After 24h, cells were washed in PBS (Gibco) and prepared for RNA extraction according to the RNeasy protocol (Qiagen), yielding around 2 μg of total RNA per sample. Illumina TruSeqTM RNA Sample preparation guide using the appropriate indices
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|
|
Library strategy |
RNA-Seq |
Library source |
transcriptomic |
Library selection |
cDNA |
Instrument model |
Illumina HiSeq 2000 |
|
|
Description |
Stimulated RNA-seq replicate 1 (S1)
|
Data processing |
An initial filtering was performed to remove reads containing residual adapter sequences (fastx_clipper, version 0.013, option –M15). Next, a quality report was generated using the FastQC software (version 0.9) checking the reads for PHRED quality (> 20), read length (> 20) and primer- contamination. Filtered reads were mapped to the human reference genome hg19 (UCSC) using TopHat (v1.3.3) with its default settings. Gene expression was measured with HT-Seq (–str=no parameter, version 0.5.3p3) using the human RefSeq annotation, release 42. DESeq was used to calculate differential expression between Nutlin3a-stimulated and non-stimulated samples. Only genes containing more then 1 read per million in at least two samples were measured. A final list of differentially expressed genes was obtained using cutoff values for both the adjusted p-value (< 0.05) and the log2 fold change (|log2FC| > 1). Genome_build: hg19 Supplementary_files_format_and_content: raw abundance counts from htseq-counts compiled in a text file and normalized counts
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|
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Submission date |
May 17, 2013 |
Last update date |
May 15, 2019 |
Contact name |
Rekin's Janky |
E-mail(s) |
rekins.janky@med.kuleuven.be
|
Organization name |
KU Leuven
|
Department |
Department of Human Genetics
|
Lab |
Laboratory of Computational Biology
|
Street address |
Herestraat 49 box 602
|
City |
Leuven |
ZIP/Postal code |
3000 |
Country |
Belgium |
|
|
Platform ID |
GPL11154 |
Series (2) |
GSE47042 |
Discovery of the p53 targetome in MCF7 cells from RNA-seq data |
GSE47043 |
Discovery of the p53 targetome in MCF7 cells from RNA-seq and ChIP-seq data |
|
Relations |
BioSample |
SAMN02146494 |
SRA |
SRX281504 |