|
| Status |
Public on Sep 21, 2023 |
| Title |
Patient_P1396_Pre_Treatment_scRNA-seq |
| Sample type |
SRA |
| |
|
| Source name |
Primary CD138+ MM cells
|
| Organism |
Homo sapiens |
| Characteristics |
patient number: P1396
time point: Pre
cell type: Primary CD138+ MM cells
library preparation: 10X Genomics Single Cell RNAseq v2 Chemistry
|
| Growth protocol |
Primary CD138+ MM cells were obtained from the BM aspirates of MM patients at the time of diagnostic procedure, using positive selection with CD138 microbeads (Miltenyi Biotech, Auburn, CA) and accordingly to manufacturer’s instructions. CD138+ fractions were subsequently analyzed by flow cytometry in order to confirm the sample’s purity. HEK293T cells were cultured in Dulbecco’s Modified Eagle’s Medium (DMEM) supplemented with 10% FBS, 100 U/mL penicillin and 100 g/mL streptomycin. All of the listed cells above were cultured at 37°C and 5% CO2.
|
| Extracted molecule |
polyA RNA |
| Extraction protocol |
Single-cell library preparation for RNA-sequencing primary MM cells were processed according to 10X Genomics reagent Kits User Guide (CG00052 v2 Chemistry). Cells were partitioned into nanoliter-scale Gel Bead-In-Emulsions (GEMs) using 10X GemCode Technology. Primers containing (i) an Illumina R1 sequence, (ii) a 16 bp 10x barcode, (iii) a 10 bp Unique Molecular Identifier (UMI) and (iv) a poly-dT primer sequence were incubated with partitioned cells resulting in barcoded, full-length cDNA from poly-adenylated mRNA. Silane magnetic beads were used to remove leftover biochemical reagents/primers, then cDNA was amplified by PCR. Enzymatic Fragmentation and Size Selection was used to optimize cDNA amplicon size prior to library construction. R1 (read 1 primer sequence) were added during GEM incubation, whereas P5, P7, a sample index (i7), and R2 (read 2 primer sequence) were added during library construction via End Repair, A-tailing, Adaptor Ligation and PCR.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic single cell |
| Library selection |
cDNA |
| Instrument model |
Illumina NextSeq 500 |
| |
|
| Data processing |
All raw FASTQs reads were aligned against the human reference genome GRCh38 with default parameters.
Assembly: Human referene genome GRCh38
Supplementary files format and content: Filtered feature - barcode HDF5 file output from CellRanger pipeline
|
| |
|
| Submission date |
Feb 13, 2023 |
| Last update date |
Sep 21, 2023 |
| Contact name |
Nizar Bahlis |
| E-mail(s) |
nbahlis@ucalgary.ca
|
| Phone |
403-220-2801
|
| Organization name |
University of Calgary
|
| Department |
Divisions of Hematology and Oncology
|
| Street address |
3330 Hospital Dr NW HMRB328
|
| City |
Calgary |
| State/province |
Alberta |
| ZIP/Postal code |
T2N4N1 |
| Country |
Canada |
| |
|
| Platform ID |
GPL18573 |
| Series (2) |
| GSE199359 |
scRNAseq Datasets Supporting Integrated Epigenetic and Transcriptional Single-Cell Analysis of t(11;14) Multiple Myeloma and Its BCL2 Dependency |
| GSE199373 |
Integrated Epigenetic and Transcriptional Single-Cell Analysis of t(11;14) Multiple Myeloma and Its BCL2 Dependency |
|
| Relations |
| BioSample |
SAMN33271091 |
| SRA |
SRX19353455 |
