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| Status |
Public on May 29, 2024 |
| Title |
WT aged sample #89 |
| Sample type |
SRA |
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| Source name |
bone marrow
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| Organism |
Mus musculus |
| Characteristics |
tissue: bone marrow
age: 12 months
cell type: Lin-cKit+
genotype: WT
index library: SIGAC5
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| Extracted molecule |
total RNA |
| Extraction protocol |
The femora, tibiae, and ilia of C57BL/6 mice of WT and STAT5ab mice were crushed and the bone marrow cells were released and the red blood cells were lysed with ammonium chloride solution (STEMCELL Technologies). The cells were stained with the fluorophore-conjugated monoclonal antibodies, and sorted using an Influx cell sorter (BD Biosciences) equipped with five lasers. The cells were sorted into 1.5 ml tubes containing PBS + 2% FCS. Sorted cells were harvested, resuspended in 0.04% BSA (Thermo Fisher) and counted. Cells were processed using the 10x Chromium™ Single Cell 3' kit (v2), following manufacturers instructions.
Libraries were prepared using 10x Chromium™ Single Cell 3' kit (v2), following manufacturers instructions. Briefly, cells were resuspended in the master mix and loaded together with partitioning oil and gel beads into the chip to generate the gel bead-in-emulsion (GEM). The poly-A mRNA from the cell lysate contained in every single GEM was retrotranscribed to cDNA, which contains an Ilumina R1 primer sequence, Unique Molecular Identifier (UMI) and the 10x Barcode. The pooled cell barcoded cDNA was then cleaned up with Silane DynaBeads, amplified by PCR and the appropriate size fragments were selected with SPRIselect reagent for subsequent library construction. During the library construction P5, P7, a sample index, and an Ilumina Read 2 are added (via End Repair, A-tailing, Adaptor Ligation, and PCR). The resulting libraries contain the P5 and P7 primer sequences required for Illumina sequencing.
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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 |
10x Genomics
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| Data processing |
The demultiplexing, barcoded processing and gene counting were made using the Cell Ranger software v2.1.1 ( https://support.10xgenomics.com/single-cell-gene-expression/software/pipelines/latest/what-is-cell-ranger)
Assembly: mm10
Supplementary files format and content: h5 compressed file with filtered UMI counts
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| Submission date |
Jan 24, 2023 |
| Last update date |
May 29, 2024 |
| Contact name |
Hugo P. Bastos |
| E-mail(s) |
hpb29@cam.ac.uk
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| Organization name |
Wellcome - MRC Cambridge Stem Cell Institute
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| Department |
Haematology
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| Street address |
Jeffrey Cheah Biomedical Centre, Puddicombe Way
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| City |
Cambridge |
| ZIP/Postal code |
CB2 0AW |
| Country |
United Kingdom |
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| Platform ID |
GPL24247 |
| Series (1) |
| GSE223632 |
Maintenance of haematopoietic stem cells by tyrosine-unphosphorylated STAT5 and JAK inhibition - STAT5 deficient HSPCs (LK) |
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| Relations |
| BioSample |
SAMN32895728 |
| SRA |
SRX19157780 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM6968282_RBG28333_filtered_gene_bc_matrices_h5.h5 |
28.5 Mb |
(ftp)(http) |
H5 |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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