|
| Status |
Public on Jan 30, 2023 |
| Title |
MCF10A_NIPBL KD_R3_RNA-seq |
| Sample type |
SRA |
| |
|
| Source name |
breast
|
| Organism |
Homo sapiens |
| Characteristics |
tissue: breast
cell line: MCF10A
cell type: epithelial
treatment: NIPBL siRNA (single oligont)
|
| Treatment protocol |
MCF10A cells were transfected with 50 nM siRNAs using DharmaFECT reagent 1 and Gibco Opti-MEM I Reduced Serum Media and harvested 72h later.
|
| Growth protocol |
MCF10A cells are grown in DMEM/F12 supplemented with 20ng/ml of EGF, 0.5mg/ml hydrocortisone, 100ng/ml of cholera toxin, 10mg/ml of insulin and 5% of horse serum.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted with total RNA (RNeasy Mini Kit, Qiagen) following manufacturer’s instructions and treated with DNaseI (Ambion).
Total RNA samples (500ng) were processed with the "NEBNext Single Cell/Low Input RNA Library Prep" kit (NEB #E6420) by following manufacturer instructions. RNA Quality scores were 9.9 on average (range 9.1-10) when assayed on a PerkinElmer LabChip analyzer. Briefly, an oligo(dT) primed reverse transcription with a template switching reaction was followed by double stranded cDNA production by limited-cycle PCR. {Non-directional} sequencing libraries were completed with the "NEBNext Ultra II FS DNA Library Prep Kit for Illumina" (NEB #E7805) and subsequently analysed on an Illumina instrument (Illumina NextSeq 550 with v2.5 reagent kits) following manufacturer's protocols.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
NextSeq 550 |
| |
|
| Data processing |
Fastq files with 86-nt single-end sequenced reads were quality-checked with FastQC and aligned to the human genome (hg19) with Nextpresso,-2.0.0 using Bowtie 0.12.7 and Samtools 0.1.16 allowing two mismatches and five multi-hits
The reads were mapped to hg19 genes using HTSeq and the differencial expression was performed using the R package DESeq2
Supplementary files format and content: txt file with raw counts
|
| |
|
| Submission date |
Jun 28, 2022 |
| Last update date |
Jan 30, 2023 |
| Contact name |
Daniel Gimenez |
| E-mail(s) |
dgimenezl@cnio.es
|
| Organization name |
CNIO
|
| Street address |
Calle de Melchor Fernández Almagro, 3
|
| City |
Madrid |
| ZIP/Postal code |
28029 |
| Country |
Spain |
| |
|
| Platform ID |
GPL21697 |
| Series (2) |
| GSE207110 |
Response of cohesin STAG1 and STAG2 to reduction of NIPBL levels [RNA-Seq] |
| GSE207116 |
Response of cohesin STAG1 and STAG2 to reduction of NIPBL levels |
|
| Relations |
| BioSample |
SAMN29402521 |
| SRA |
SRX15927957 |
