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| Status |
Public on Mar 08, 2023 |
| Title |
Mc1-IRAP-LIB |
| Sample type |
SRA |
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| Source name |
monoocyte derived macrophage
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| Organism |
Equus caballus |
| Characteristics |
cell type: monoocyte derived macrophage
treatment: ACS
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| Treatment protocol |
Equine monocyte-derived macrophages were stimulated with IL-1ß (10 ng/mL) and OTs (MSC-CM, PRP lysate, ACS) were added at the same time to macrophage culture media at a ratio of 1:3 OT to complete growth media in culture (i.e., 25% OT in culture media) Following transient addition of treatments for 24 hours in culture, macrophages were washed three times with phosphate buffered saline (PBS) , collected in RNA lysis buffer (350 L/sample) and frozen at -20C until RNA isolation was performed.
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| Growth protocol |
To generate macrophage cultures, equine peripheral blood mononuclear cells were isolated from whole blood of two horses by density gradient centrifugation (Ficoll- Paque TM plus, GE Healthcare Bio-Sciences) and cultured in macrophage media (Dulbecco’s Modified Eagle’s Medium supplemented with 10% fetal bovine serum, non-essential amino acids, and penicillin/streptomycin antibiotics; SigmaAldrich) with human M-CSF (PeproTech, Rocky Hill, NJ USA 80553) at 30 ng/mL to stimulate differentiation into macrophages in five days, as previously described [332].
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| Extracted molecule |
total RNA |
| Extraction protocol |
Qiagen Rneasy
mRNA was enriched using oligo(dT) beads, followed by cDNA library generation using TruSeq RNA Library Prep Kit
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina NovaSeq 6000 |
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| Description |
F_IRAP
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| Data processing |
demultiplexed Fasq reads generated from Novogene were analyzed using Partek® Flow® software, v10.0
Reads were trimmed for Phred score of 20, adapters removed using cutadapt
Trimmed reads were aligned using STAR 2.7.3 using and annotated with Ensembl EquCab3.0.107. Feature counts were generated with Htseq
Differential analysis was computed using counts normalized to CPM, using DESeq
Pathway analysis was performed with GSEA (Gene Set Enrichment Analysis) v4.2.1using Hallmark pathways
Assembly: EquCab3.0
Supplementary files format and content: txt file of cpm normalized counts filtered for protein coding genes
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| Submission date |
Feb 02, 2023 |
| Last update date |
Mar 08, 2023 |
| Contact name |
Steven Dow |
| E-mail(s) |
sdow@colostate.edu
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| Organization name |
Colorado State University
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| Department |
Clinical Sciences
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| Lab |
ACC 254
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| Street address |
300 W Drake Rd
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| City |
Fort Collins |
| State/province |
CO |
| ZIP/Postal code |
80524 |
| Country |
USA |
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| Platform ID |
GPL26749 |
| Series (1) |
| GSE224326 |
Distinct differences in immunological properties of equine orthobiologics revealed by functional and transcriptomic analysis using an activated macrophage readout system |
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| Relations |
| BioSample |
SAMN33017632 |
| SRA |
SRX19259106 |
| 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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