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GEO help: Mouse over screen elements for information. |
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| Status |
Public on Nov 02, 2022 |
| Title |
Fresh_Lin_Depleted_scRNA-seq |
| Sample type |
SRA |
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| Source name |
Bone marrow
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| Organism |
Mus musculus |
| Characteristics |
cell type: Freshly isolated Lin-, c-Kit+ HSPCs
tissue: Bone marrow
strain: BL/6
age: 6-8 weeks old
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| Extracted molecule |
total RNA |
| Extraction protocol |
For scRNA-seq extract protocol, For C57BL/6 Lineage-depleted BM cells were seeded at 1E3 cells per 35 mm dish in StemCell Technologies’ MethocultTM GF M3434 methylcellulose media and cultured for two weeks with either 30 uM SB204990 (Tocris Cat# 4962) or an equivalent volume of DMSO. Cells were harvested and resuspended in PBS alongside freshly isolated lineage-depleted BM cells from a C57BL/6 mouse. Cells were then stained for viability using eBioscience™ Fixable Viability Dye eFluor™ 780 (Thermo Fisher Scientific 65-0865-14) and flow-sorted for viable cells. For scATAC-seq extract protocol, C57BL/6 Lineage-depleted BM cells were seeded at 1E3 cells per 35 mm dish in StemCell Technologies’ MethocultTM GF M3434 methylcellulose media and cultured for two weeks with either 30 uM SB204990 (Tocris 4962) or an equivalent volume of DMSO. Cells were harvested and resuspended in PBS alongside freshly isolated lineage-depleted BM cells from a C57BL/6 mouse.
For scRNA-seq library construction, Cells were submitted to Vanderbilt University’s VANTAGE Core, where they were prepared for single-cell 5’ RNA sequencing using the 10x Genomics Chromium system. Libraries were prepared using P/N 1000014, 1000020, 1000080, and 120262 according to the manufacturer’s protocol. The libraries were sequenced using the NovaSeq 6000 with 150 bp paired-end reads. RTA (version 2.4.11; Illumina) was used for base calling. For scATAC-seq library construction, Nuclei were isolated and submitted to Vanderbilt University’s VANTAGE Core, where they were prepared for scATAC-seq following the manufacturer’s protocol using P/N 1000111, 1000086, and 1000084.
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic single cell |
| Library selection |
cDNA |
| Instrument model |
Illumina NovaSeq 6000 |
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| Description |
FLD_RNA, replicate 1, scRNAseq
10x Genomics
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| Data processing |
For scRNA-seq, The demultiplexing, barcode processing, gene counting and aggregation were made using the Cell Ranger software v3.0.2 (https://support.10xgenomics.com/single-cell-gene-expression/software/pipelines/latest/what-is-cell-ranger). For scATAC-seq, demultiplexing, barcode processing, peak calling, and aggregation were made using the Cell Ranger ATAC software v1.2.0. Files were imported into R using ArchR v1.0.0 (filterTSS = 8, filterFrags = 2819), with filterFrags value set based on the distribution of transcription start site (TSS) vs. the log transform of unique fragments generated by ArchR.
Assembly: mm10
Supplementary files format and content: Tab-separated values files, bed files for scATAC-seq, and matrix files
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| Submission date |
Nov 02, 2022 |
| Last update date |
Nov 03, 2022 |
| Contact name |
Dalton Lee Greenwood |
| E-mail(s) |
dalton.l.greenwood@vanderbilt.edu
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| Organization name |
Vanderbilt University
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| Department |
PMI
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| Lab |
Jeff Rathmell Lab
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| Street address |
2201 West End Ave
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| City |
Nashville |
| State/province |
TN |
| ZIP/Postal code |
37235 |
| Country |
USA |
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| Platform ID |
GPL24247 |
| Series (1) |
| GSE217080 |
Gene expression profile at single cell level of hematopoietic stem and progenitor cells (HSPCs) from mouse bone marrow cultured in M3434 StemCell methylcellulose media |
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| Relations |
| BioSample |
SAMN31572153 |
| SRA |
SRX18120636 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM6704896_FLD_RNA_barcodes.tsv.gz |
3.5 Kb |
(ftp)(http) |
TSV |
| GSM6704896_FLD_RNA_features.tsv.gz |
244.7 Kb |
(ftp)(http) |
TSV |
| GSM6704896_FLD_RNA_matrix.mtx.gz |
8.8 Mb |
(ftp)(http) |
MTX |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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