|
| Status |
Public on Mar 09, 2022 |
| Title |
T3-2 single-cells |
| Sample type |
SRA |
| |
|
| Source name |
Induced tenocytes
|
| Organism |
Mus musculus |
| Characteristics |
cell types: ScxGFP iPSCs derived tendon progenitors
stimulation: TGF-β2 for 3 days
|
| Treatment protocol |
Mesodermal cells (T0) were induced from ScxGFP iPSCs in DF (DMEM-F12) supplemented with 5% FBS, 2 mM L-glutamate, and ITS (10 ng/ml insulin, 10 ng/ml transferrin, 3 × 10^-8 M sodium selenite) for 4 days. T0 cells were cutured in DF supplemented with 1% FBS, 2 mM L-glutamate, ITS, and 10 ng/ml recombinant human TGF-β2 for 3, 6, or, 9 days (T3, T6, T9).
|
| Growth protocol |
ScxGFP iPSCs were cultured on feeder cells under 2i LIF condition.
|
| Extracted molecule |
polyA RNA |
| Extraction protocol |
Cells were sorted and lysed in 1ul of lysis buffer containing (0.3% NP40, 0.12mM dNTPs, 1U RNase Inhibitor, 0.11 uM 384-well-unique Reverse Transcribe primer).
CEL-Seq2 protocol was used for single cell analysis with some modifications that SuperScripII enzyme to MaximaH minus (Thermo Scientific, EP0752) and 2nd strand buffers to 2nd Strand Module (NEB,E6111). 384 cells in the same plate were pooled together after reverse transcription.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 1500 |
| |
|
| Description |
R2(36bp) is insert reads and R1(15bp) is UMI(6bp) + cell barcode(9bp) reads for 384 cells. Please use 384_CellBarcode_list.txt.gz for demultiplexing.
|
| Data processing |
Unique-molecular-identifiers and cell baecodes in the read1 were extracted using UMI-tools (version 1.0.1) with the command: umi_tools extract --bc-pattern=NNNNNNCCCCCCCCC --stdin Read1.fastq --read2-in Read2.fastq --read2-stdout --filter-cell-barcode --whitelist=384_usedCB.txt --quality-encoding phred33 --quality-filter-mask 20. Reads were aligned to GRCm38 by HISAT2 (version 2.2.1). UMI count matrix was obtained by featureCounts (version v2.0.1) and UMI-tools with the command: umi_tools count --method=unique --per-gene --per-cell --gene-tag=XT. The unique molecular identifiers duplicates were discarded.
Genome_build: GRCm38
Supplementary_files_format_and_content: raw count matrix:(CSV) and cell barcode list(txt), gzipped
|
| |
|
| Submission date |
Mar 08, 2021 |
| Last update date |
Mar 09, 2022 |
| Contact name |
Yasuyuki Ohkawa |
| E-mail(s) |
[email protected]
|
| Organization name |
Medical Institute of Bioregulation
|
| Lab |
Division of Transcriptomics
|
| Street address |
3-1-1 Maidashi
|
| City |
Fukuoka |
| ZIP/Postal code |
8128582 |
| Country |
Japan |
| |
|
| Platform ID |
GPL18480 |
| Series (1) |
| GSE168451 |
Tenogenic induction from induced pluripotent stem cells unveils the trajectory towards tenocyte differentiation. |
|
| Relations |
| BioSample |
SAMN18205475 |
| SRA |
SRX10265230 |
