|
| Status |
Public on Nov 27, 2019 |
| Title |
RNAExp31 |
| Sample type |
SRA |
| |
|
| Source name |
Jurkat cell line and Nalm6 cell line
|
| Organism |
Homo sapiens |
| Characteristics |
cell line: Jurkat cell line and Nalm6 cell line
molecule type: various targeted RNA transcripts
|
| Growth protocol |
Jurkat cells and Nalm-6 cells were obtained from America Type Tissue Collection (ATCC). Cells at a density of ~1×106/mL were fixed in RPMI medium without serum in 1.6% paraformaldehyde (Electronic Microscopy Sciences) for 10 min at room temperature under gentle agitation. Cells were pelleted and permeabilized with ice-cold methanol for at least 10 min on ice. Once in methanol cells can be stored at −80 °C for several weeks without loss of antibody signal or RNA degradation.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
QIAquick Gel Extraction Kit (Qiagen, 28704).
PCR with Illumina dual index primers
|
| |
|
| Library strategy |
OTHER |
| Library source |
transcriptomic |
| Library selection |
other |
| Instrument model |
Illumina NextSeq 500 |
| |
|
| Data processing |
Library strategy: Single cell barcoding by split-pool synthesis via ligation
FLASH-1.2.11 was used to merge the paired end reads.
seqtk-v1.2 was used to convert merged fastq file to fasta format.
The reads were then parsed to identify cell-id and marker-id based on the white-list info in the associated qdata folder, deduplicated based on cell-id and marker-ids and finally chimeric reads were filtered out.
Expression counts were then used to remove putative debris and doublets cells based on the criteria listed in Singlet_settings.txt in the corresponding qdata folder.
Expression counts matrix was then normalized by scaling the counts by row-means and multiplying by a factor of 10.
A github repository for the software used to process the raw-fastqs is available at the following location: https://github.com/bioinform/QBC_Single_Cell_Analysis_NGS
Supplementary_files_format_and_content: Tab separated text files containing jittered expression counts (both normalized and un-normalized) for different markers for each individual cell
The *tar.gz contains various files containing white-list information for cell-barcode ids, anchor-sequence info, marker-sequence info and settings information for removal of cell-debris and cell-doublets. The files in this folder is used to process the raw fastqs for each of the experiment.
|
| |
|
| Submission date |
Nov 26, 2019 |
| Last update date |
Nov 27, 2019 |
| Contact name |
Maeve Ohuallachain |
| E-mail(s) |
maeve.ohuallachain@roche.com
|
| Organization name |
ROCHE SEQUENCING SOLUTIONS INC
|
| Street address |
4300 Hacienda Drive
|
| City |
Pleasanton |
| State/province |
CALIFORNIA |
| ZIP/Postal code |
94588 |
| Country |
USA |
| |
|
| Platform ID |
GPL18573 |
| Series (1) |
| GSE130784 |
Ultra-High Throughput Single Cell Analysis of Proteins and RNAs by Split-pool Synthesis |
|
| Relations |
| BioSample |
SAMN13390045 |
| SRA |
SRX7217831 |
