|
| Status |
Public on Jun 24, 2018 |
| Title |
Primary breast cancer 1 CD103- |
| Sample type |
SRA |
| |
|
| Source name |
Primary Tumor
|
| Organism |
Homo sapiens |
| Characteristics |
breast cancer subtype: triple negative
phenotype: CD45+MHCI+CD3+CD8+CD69+CD103-
tissue: Primary breast cancer 1
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Single cell suspensions of breast cancer tissue were obtained by enzymatic dissociation on the day of surgery, from one primary TNBC, one primary ER negative HER2 amplified breast cancer, and one TNBC liver metastasis. Viable cells were FACS sorted on BD FACSAria II Cell Sorter (BD Biosciences) by propidium iodide staining and further sorted for CD8+CD69+CD103+ and CD8+CD69+CD103- TIL subsets. Total RNA was extracted using the RNAqueous-4PCR RNA isolation kit (Thermo Fisher) with cell concentrations ranging between 1 x 10^5 - 2 x 10^5 cells for RT PCR. Between 50 and 200ng total RNA were used per sample.
Libraries were prepared using the Illumina Trussed RNA Sample Prep Kit v2 following the vendors protocol. Briefly, poly-A mRNA was purified using poly-T magnetic beads, fragmented using divalent cations under elevated temperature and reverse transcribed to cDNA with random primers. Indexed adaptors were then ligated and the libraries amplified. 15 to 19 cycles of PCR were necessary. Library concentration was measured using Qubit High Sensitivity kit and fluorometer (Thermo Fisher Scientific) and the quality checked using the High Sensitivity D1000 reagents in an Agilent Tape Station.
polyA selected unstranded RNASeq
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina NextSeq 500 |
| |
|
| Description |
179-T2
Raw data (controlled access) at EGAN00001668304
|
| Data processing |
RNASeq data were quantified using kallisto (0.43.0), against the Ensembl transcript reference (release 79, GRCh38)
Count matrix was constructed using tximport (1.4.0) with countsFromAbundance = "lengthScaledTPM"
Genome_build: GRCh38
Supplementary_files_format_and_content: Gene counts for 6 samples, tsv
|
| |
|
| Submission date |
Feb 21, 2018 |
| Last update date |
Jun 25, 2018 |
| Contact name |
Chengzhong Ye |
| E-mail(s) |
ye.c@wehi.edu.au
|
| Organization name |
Walter and Eliza Hall Institute of Medical Research
|
| Department |
Bioinformatics Division
|
| Lab |
Speed Lab
|
| Street address |
Walter and Eliza Hall Institute of Medical Research, 1G Royal Parade
|
| City |
Parkville |
| State/province |
Victoria |
| ZIP/Postal code |
3052 |
| Country |
Australia |
| |
|
| Platform ID |
GPL18573 |
| Series (1) |
| GSE110938 |
Differential gene expression of tissue-resident memory and effector memory T cells in breast cancer |
|
| Relations |
| BioSample |
SAMN08578670 |
