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| Status |
Public on Apr 30, 2021 |
| Title |
Epithelial progenitors E14.5 |
| Sample type |
SRA |
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| Source name |
Trachea epithelial progenitors
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| Organism |
Mus musculus |
| Characteristics |
strain: C57BL/6
developmental stage: E14.5
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| Treatment protocol |
None
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| Growth protocol |
None
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| Extracted molecule |
polyA RNA |
| Extraction protocol |
To prepare single-cell suspensions of epithelial cells in 7 time points (E12.5, E13.5, E14.5, E16.5, E18.5, and 2months), the epithelial sheet was peeled off from mesenchymal tissue in developing trachea with a tungsten needle after the incubation with 175U/ml collagenase typeⅠ (Worthington Biochemical Corporation, CLS1) at 37℃ for 6 - 120 min. Single-cell suspensions were made through the incubation with Trypsin-EDTA (0.25%) (Thermo Fisher Scientific, 25200056) at 37℃ for 15 min, then loaded onto Chromium Single Cell A Chips (10X Genomics, PN-1000009) for the Chromium Single Cell 3′ Library v2 (10X Genomics, PN-120233) according to the manufacturer’s recommendations (10X Genomics)
Signle-cell suspensions were loaded onto Chromium Single Cell A Chips (10X Genomics, PN-1000009) for the Chromium Single Cell 3′ Library v2 (10X Genomics, PN-120233) according to the manufacturer’s recommendations (10X Genomics). Briefly, single-cell gel bead-in-emulsions (GEMs) were generated from loaded cell suspensions by a Chromium Controller instrument (10X Genomics). After performing GEM-reverse transcriptions (GEM-RTs), GEMs were harvested and the cDNAs were amplified and cleaned up with SPRIselect Reagent Kit (Beckman Coulter, B23318). Indexed sequencing libraries were constructed using Chromium Single Cell 3′ Library v2 (10X Genomics, PN-120233) for enzymatic fragmentation, end-repair, A-tailing, adaptor ligation, ligation cleanup, sample index PCR, and PCR cleanup. Libraries were sequenced on a HiSeq1500 (Illumina) to obtain a sequencing depth of around 50,000 reads per cells.
single cell RNA-seq using Chromium system (PolyA+ RNA)
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 1500 |
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| Description |
E12-E18_barcodes.tsv
E12-E18_matrix.mtx
E12-E18_genes.tsv
E12-18_Seurat_Processed.Robj
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| Data processing |
Basecalls performed using Illumina RTA 1.18.64 and the fastq files were generated by Cellranger v2.2.0 and bcl2fastq 2.19.0.316
signle cell RNA-seq reads were aligned to the mm10 genome reference using Cellranger v2.2.0 with default parameters
Seurat v2.3.4: the cells meeting any of the following criteria were omitted from further analyses for the quality control; <1,000 or >5,000 UMIs, > 7.5% of reads mapping to mitochondria genes, or EpCAM negative cells. For clustering, principal-component analysis was performed for dimension reduction. Top 15 principal components (PCs) were selected by using a permutation-based test implemented in Seurat and passed to t-Distributed Stochastic Neighbor Embedding (tSNE) for clustering visualization. To maintain a standard procedure for clustering, a value of 0.8 for the resolution was used
Genome_build: mm10
Supplementary_files_format_and_content: Filtered gene-barcode matrix of CellRanger output including sparse matrix (MEX), cellular barcodes (TSV), and gene list (TSV). Processed data with Seurat (Robj)
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| Submission date |
Jun 17, 2020 |
| Last update date |
Apr 30, 2021 |
| Contact name |
Hirofumi Kiyokawa |
| E-mail(s) |
h.kiyokawa@gmail.com
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| Phone |
81783063199
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| Organization name |
Riken
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| Department |
Center for Biosystems Dynamics Research
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| Lab |
Laboratory for lung development and regenration
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| Street address |
2-2-3 Minatojima-minamimachi, Chuo-ku
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| City |
Kobe |
| State/province |
Hyogo |
| ZIP/Postal code |
6500047 |
| Country |
Japan |
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| Platform ID |
GPL18480 |
| Series (1) |
| GSE152692 |
Comprehensive single cell RNA-seq datasets of mice trachea epithelial progenitors in 6 time points from E12.5 to E18.5 and mature epithelial cells in 2-month mice |
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| Relations |
| BioSample |
SAMN15300201 |
| SRA |
SRX8567942 |
| Supplementary data files not provided |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data are available on Series record |
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