|
| Status |
Public on May 20, 2024 |
| Title |
Human Prostate Biopsy, HMP22 |
| Sample type |
SRA |
| |
|
| Source name |
Prostate tumor, lymph node
|
| Organism |
Homo sapiens |
| Characteristics |
tissue: Prostate tumor, lymph node
|
| Extracted molecule |
polyA RNA |
| Extraction protocol |
Fresh benign and malignant tissue was mechanically cut using a scalpel into small pieces (~1–5 mm3). The tissue was then processed and dissociated in 5–10 mg/ml collagenase type II (Gibco) solution in adDMEM/F12+/+/+ with 10 µM Y–27632 dihydrochloride for 30 minutes to 2 hours on a 37oC shaking platform. This was followed by a 1 minute 0.5 M EDTA wash at room temperature, and subsequent digestion with TrypLE (Gibco) with 10 µM Y–27632 dihydrochloride for 5–10 minutes at 37oC on a shaking platform until a single cell suspension was obtained. If more than 10% of doublets were present (visual inspection) or there was evidence for <80% viability (hemocytometer, using 0.2% Trypan Blue), then cells were FACS–sorted for singlets and viability using a DAPI. Please refer to methods of paper for further details.
Dissociated cells were subjected to scRNA–seq using 10X genomics Chromium Single Cell 3’ Library and Gel bead Kit (v3 for all human biospies, except v2 for HP95T as previously described PMID: 32355025) per manufacturer’s protocol. ~3000 to 10,000 cells per sample was encapsulated and barcoded following the manual. Sample viability varied between 72 and 95% (0.2% Trypan Blue). The final sequencing libraries were double–size purified (0.6–0.8X) with SPRI beads and sequenced on Illumina Nova–Seq platform (R1–26 cycles, i7–8 cycles, R2–70 cycles or higher).
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic single cell |
| Library selection |
cDNA |
| Instrument model |
Illumina NovaSeq 6000 |
| |
|
| Description |
10x Genomics
|
| Data processing |
SEQC alignment
Cell filtering with Seurat (percent.mt < 25, nCount > 200)
DoubletDetection for doublet detection
log normalization
Cell cycle score regression with Seurat
Data scaling with Seurat
Seurat for downstream analysis
Assembly: hg38
Supplementary files format and content: count matrix (.mtx.gz)
Supplementary files format and content: feature and barcode (.tsv.gz)
Supplementary files format and content: seurat object for whole cells and tumor cells (.rds)
|
| |
|
| Submission date |
Apr 22, 2024 |
| Last update date |
May 20, 2024 |
| Contact name |
Samir Zaidi |
| E-mail(s) |
zaidis@mskcc.org
|
| Organization name |
Memorial Sloan Kettering Cancer Center
|
| Street address |
1275 York Avenue
|
| City |
New York |
| ZIP/Postal code |
10065 |
| Country |
USA |
| |
|
| Platform ID |
GPL24676 |
| Series (1) |
| GSE264573 |
Single Cell Analysis of Treatment–Resistant Prostate Cancer: Implications of Cell State Changes for Cell Surface Antigen Targeted Therapies |
|
| Relations |
| BioSample |
SAMN41047769 |
| SRA |
SRX24336216 |
