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GEO help: Mouse over screen elements for information. |
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| Status |
Public on Aug 26, 2016 |
| Title |
GMP cell R3.58 |
| Sample type |
SRA |
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| Source name |
Bone Marrow
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| Organism |
Mus musculus |
| Characteristics |
cell type: GMP Progenitors
strain: C57BL/6
cdna synthesis: Fluidigm C1 protocol
library prep: Nextera XT DNA Library Preparation Kit (Illumina Inc, Santa Clara, CA)
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| Extracted molecule |
total RNA |
| Extraction protocol |
Cells were harvested from bone marrow. Femurs, tibias and iliac crest were harvested immediately after euthanasia and put in cold PBS+2%FBS. Bones were crushed with mortar and pestle, filtered and washed in cold PBS+2%FBS, then enriched using CD117 MicroBeads on a Automacs Pro separator (Miltenyi, San Diego, CA). CD117+ cells were stained with CD16/32-PerCp-ef710 (clone 93), CD3-biotin (clone 145-2C11), CD4-biotin (clone RM4-5), CD8-biotin (clone 53-6.7), CD11b-biotin (clone M1/70), CD19-biotin (clone 6D5), Gr1-biotin (clone RB6-8C5), Ter119-biotin (clone Ter-119) CD45R-biotin (clone RA3-6B2), CD117-APC (clone 2B8), Sca-1-Pe-Cy7 (clone D7), CD34-BV421 (clone RAM34), CD115-BV605 (clone TR15-12F1 2.2), CD135-PE (A2F10.1), CCR3-FITC (clone 83101), SiglecF (clone E50-2440) and Gr1-FITC (clone RB6-8C5). Cell sorting was performed on MoFloXDP (Beckman Coulter, Brea, CA) or BD FACSAria II with a 100μm nozzleand subjected to flow cytrometric sorting of Lin- CD117+ Sca1- CD16/32+ CD34+ cells (GMP).
cDNA was made with Clontech SMARTer® Kit “the mRNA Seq: RT + Amp (1771x), Libraries were constructed using Nextera XT DNA Library Preparation Kit
Single cells were separated using Fluidigm C1 system
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 2500 |
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| Description |
Lin- CD117+ Sca1- CD16/32+ CD34+
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| Data processing |
RNA reads were aligned to the reference mm9 mouse genome using Bowtie2.
Single-cell and matched bulk sorted RNA-Seq were analyzed using RSEM to estimate transcripts per million mapped reads (TPM) for all genes.
Differentially expressed genes were identified using AltAnalyze using a FDR adjusted empirical Bayes moderated t-test p<0.05.
Genome_build: mm9
Supplementary_files_format_and_content: Tab-delimited text file of log2 (incremented by 1 before log2) TPM RSEM expression values
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| Submission date |
Jun 24, 2015 |
| Last update date |
May 15, 2019 |
| Contact name |
H. Leighton Grimes |
| E-mail(s) |
Lee.Grimes@cchmc.org
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| Phone |
513-636-6089
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| Organization name |
Cincinnati Childrens Hospital Medical Center
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| Department |
Immunobiology
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| Lab |
Grimes
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| Street address |
3333 Burnet Ave. MLC 7038
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| City |
Cincinnati |
| State/province |
OH |
| ZIP/Postal code |
45229 |
| Country |
USA |
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| Platform ID |
GPL17021 |
| Series (2) |
| GSE70240 |
Using single-cell RNA-Seq for unbiased analysis of developmental hierarchies (single cell RNA Seq of GMP) |
| GSE70245 |
Using single-cell RNA-Seq for unbiased analysis of developmental hierarchies |
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| Relations |
| BioSample |
SAMN03792572 |
| SRA |
SRX1072670 |
| 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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