|
| Status |
Public on May 31, 2024 |
| Title |
SW480_spheres_rep2 |
| Sample type |
SRA |
| |
|
| Source name |
colon
|
| Organism |
Homo sapiens |
| Characteristics |
tissue: colon
cell line: SW480
cell type: grade 3-4 colon adenocarcinoma
subpopulation: sphere subpopulation
|
| Treatment protocol |
no treatment was applied
|
| Growth protocol |
SW480 subpopulations were sorted and grown in standard culture medium for 4 days before RNA was extracted.
|
| Extracted molecule |
polyA RNA |
| Extraction protocol |
RNA was harvested using Trizol reagent.
RNA-Seq was performed at the Erasmus MC Center for Biomics (Rotterdam, The Netherlands) according to manufacturer’s instructions (Illumina) and using the TruSeq RNA sample prep kit v2. Briefly, polyA containing mRNA molecules were purified using poly-T oligo attached magnetic beads. Following purification, the mRNA was fragmented into ~200 bp fragments using divalent cations under elevated temperature. The cleaved RNA fragments were copied into first strand cDNA using reverse transcriptase and random primers. This was followed by second strand synthesis using DNA polymerase I and RNaseH treatment. These cDNA fragments were end repaired, a single A base was added and Illumina adapters were ligated. The products were purified and size selected on gel and enriched by PCR. The PCR products were purified by Qiaquick PCR purification and used for cluster generation according to the Illumina cluster generation protocols.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 2000 |
| |
|
| Data processing |
Illumina paired-end reads of 76 bp were trimmed by removing the TrueSeq adapter sequences using Trimmomatic (v.0.33).
Reads were mapped to the human reference genome build hg38 with the RNA-seq aligner STAR (v2.4.2a) and the Homo sapiens GENCODE v23 annotation.
Raw counts were summed with featureCounts (v2.0.3)
Downstream analysis was done in R with DESeq2 (1.36.0).
Counts were normalized with a variance stabilizing transformation (VST).
Assembly: hg38
Supplementary files format and content: Excel file includes Ensemble Gene Ids and variance stabilizing transformation (VST) normalized counts.
|
| |
|
| Submission date |
Jan 12, 2024 |
| Last update date |
May 31, 2024 |
| Contact name |
Mathijs Verhagen |
| E-mail(s) |
m.p.verhagen@erasmusmc.nl
|
| Organization name |
Erasmus Medical Center
|
| Street address |
Doctor Molewaterplein 40
|
| City |
Rotterdam |
| ZIP/Postal code |
3015 GD |
| Country |
Netherlands |
| |
|
| Platform ID |
GPL11154 |
| Series (1) |
| GSE253110 |
The SW480 cell line as a model of resident and migrating colon cancer stem cells |
|
| Relations |
| BioSample |
SAMN39423186 |
| SRA |
SRX23203565 |
