|
| Status |
Public on Dec 22, 2015 |
| Title |
CD8_93 scRNA-seq |
| Sample type |
SRA |
| |
|
| Source name |
Single CD8+ cells grown for multiple generations on-chip after 30h activation in vitro with anti-CD3/28
|
| Organism |
Mus musculus |
| Characteristics |
cell type: Activated CD8+ T cells
strain: C57BL/6J
hours since division: 6.6
sister: CD8_94
cousin 1: CD8_91
cousin 2: CD8_92
doublet?: No
|
| Treatment protocol |
The CD8+ T cells were activated with 5 μg/mL plate-bound anti-mouse CD3 (clone: 145-2C11, BioLegend catalog number 100314) and 2 μg/mL of anti-mouse CD28 (clone: 37.51, BioLegend catalog number 102112) in solution for 30h prior to loading in to the device.
|
| Growth protocol |
L1210 cells were cultured in the device in RPMI 1640 (Gibco) with 10% FBS and 1% penicillin-streptomycin solution (Gibco). CD8+ T cells were cultured in the device in RPMI 1640 (Gibco) with 10% FBS, 55μM 2-mercaptoethanol (Gibco), 1% penicillin-streptomycin solution (Gibco) and 100 U/mL IL2 (PeproTech) after 30h activation in vitro (see below).
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Single cells were flushed from the device with 5 ul of PBS directly in to 5 ul of 2X TCL lysis buffer (Qiagen) resulting in a total volume of 10 ul of single-cell lysate. These samples were immediately frozen on dry ice and subsequently stored at -80°C prior to library preperation and sequencing.
Libraries were constructed using the Smart-Seq2 protocol for single cells.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina NextSeq 500 |
| |
|
| Description |
CD8 scRNA-seq
|
| Data processing |
Illumina bcl2fastq2.15.0.4 was used to generat .fastq files from .bcl files.
tophat2 was used to align the scRNA-seq data
rsem-1.2.3 generated an expression matrix in transcripts per million.
mm10 ucsc genomestudio
Genome_build: mm10
Supplementary_files_format_and_content: RSEM transcripts per million matrix with genes as rows and samples as columns.
|
| |
|
| Submission date |
Nov 12, 2015 |
| Last update date |
May 15, 2019 |
| Contact name |
Robert Kimmerling |
| E-mail(s) |
rjkimmer@mit.edu
|
| Phone |
6318793159
|
| Organization name |
MIT
|
| Department |
Biological Engineering
|
| Lab |
Manalis
|
| Street address |
32 Vassar St., 76-221
|
| City |
Cambridge |
| State/province |
Massachusetts |
| ZIP/Postal code |
02139 |
| Country |
USA |
| |
|
| Platform ID |
GPL19057 |
| Series (1) |
| GSE74923 |
A microfluidic platform enabling single cell RNA-seq of multigenerational lineages |
|
| Relations |
| BioSample |
SAMN04262015 |
| SRA |
SRX1427341 |
