|
| Status |
Public on Jan 21, 2019 |
| Title |
Mm_ErbB2_msphr_Nut3_rna_rep3 |
| Sample type |
SRA |
| |
|
| Source name |
Mammosphere culture
|
| Organism |
Mus musculus |
| Characteristics |
strain: FVB/Hsd
tissue: breast tumor
age: 12-15 weeks old
genotype: transgenic MMTV-ErbB2
infection: not infected
treatment: Nutlin-3 (2.5 uM for 72h)
passage: M6-M7 (72 hours after plating)
|
| Growth protocol |
cells were isolated from mammary tissue derived from WT or transgenic mice and plated in anchorage independent condition (polyHEMA coated plates) to form mammospheres. Mammospheres were serially passaged in STEM Medium (serum-free mammary epithelial basal medium MEBM, supplemented with 2 mM glutamine, 100 U/ml penicillin and 100 μg/ml streptomycin, 5 μg/ml insulin, 0.5 μg/ml hydrocortisone, 2% B27, 20 ng/ml EGF and βFGF and 4 μg/ml heparin). Mammospheres were disaggregated and RNA extracted.
|
| Extracted molecule |
polyA RNA |
| Extraction protocol |
RNA from mammospheres was extracted with Maxwell® 16 LEV simplyRNA cells kit (Promega) and reverse transcribed using random primers and ImProm-II™ reverse transcriptase (Promega), following manufacturer’s instructions.
Libraries of template RNA molecules were prepared from 0.5 to 2 μg high quality input RNA using the Illumina® TruSeq® RNA Sample Preparation Kit v2, following manufacturer’s instructions.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 2000 |
| |
|
| Data processing |
Basecalls performed using CASAVA version 1.9
Data were filtered using the Illumina quality control flag
Reads were aligned to the mus musculs genome build mm10 using tophat (v2.0.8), which uses Bowtie (2.1.0) and Samtools (v0.1.18) using default parameters and --transcriptome Mus Musculus index, UCSC mm10 gtf.
Raw read counts were summarised using HTSeq (v0.5.3p9), then normalized for library size differences using estimateSizeFactors function from DESeq2 package (v1.10.1).
Genome_build: mm10
Supplementary_files_format_and_content: tab-delimited text files include normalized expression value, log2fold change, pvalue and adjusted pvalue for each gene for each DE analysis
|
| |
|
| Submission date |
Sep 16, 2016 |
| Last update date |
May 15, 2019 |
| Contact name |
Pier Giuseppe Pelicci |
| Organization name |
European Institute of Oncology
|
| Department |
Department of Experimental Oncology
|
| Street address |
Via Adamello, 16
|
| City |
Milano |
| ZIP/Postal code |
20139 |
| Country |
Italy |
| |
|
| Platform ID |
GPL13112 |
| Series (2) |
| GSE87003 |
Myc activation in breast cancer is due to p53-loss and sustains self-renewal of cancer stem cells (RNA-Seq) |
| GSE87004 |
Myc activation in breast cancer is due to p53-loss and sustains self-renewal of cancer stem cells |
|
| Relations |
| BioSample |
SAMN05781427 |
| SRA |
SRX2170304 |
