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Status |
Public on Dec 15, 2023 |
Title |
c1829_STIM_canine_PHA |
Sample type |
SRA |
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Source name |
Blood
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Organism |
Canis lupus familiaris |
Characteristics |
tissue: Blood cell line: primary tissue cell type: PBMC treatment: PHA
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Treatment protocol |
phytohemagglutinin (PHA) (Sigma-Aldrich, St. Louis, MO) was added to cultures at a concentration of 5 ug/ml
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Growth protocol |
Following Ficoll separation, PBMC for both species were washed with PBS, counted and added to 24-well plates in triplicate cultures, at a cell density of 2 X 10^6 cells per mL in complete culture medium. Cell culture medium consisted of DMEM supplemented with amino acids (essential and non-essential), 1% penicillin-streptomycin, Glutamax (ThermoFisher Scientific, Waltham, MA) and 10% fetal bovine serum (Peak Serum, Inc, Wellington, CO). For T cell activation, phytohemagglutinin (PHA) (Sigma-Aldrich, St. Louis, MO) was added to cultures at a concentration of 5 ug/ml for 24h (RNA sequencing).
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Extracted molecule |
total RNA |
Extraction protocol |
RNA was extracted from cells using Qiagen RNeasy mini kit the mRNA was purified using poly-T oligo-attached magnetic beads. After fragmentation, the first strand cDNA was synthesized using random hexamer primers followed by the second strand cDNA synthesis. The library was completed following end repair, A-tailing, adapter ligation, size selection, amplification, and purification.
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Library strategy |
RNA-Seq |
Library source |
transcriptomic |
Library selection |
cDNA |
Instrument model |
Illumina NovaSeq 6000 |
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Description |
c1829_STIM Partek_Canine_PBMC-_PHA_Normalized.txt Partek_dog_vs_human_combined_normalized_ct.txt
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Data processing |
Partek Flow software, version 10.0 Raw data were filtered by removing reads containing adapters and reads containing N > 10% and for Phred scores >30 Filtered reads were aligned with STAR 2.7.3a, using either CanFam3.1 genome assembly or GRCh38 human genome assembly Aligned reads were annotated and counted using HT-seq with Ensembl 104 for CanFam3.1 and release 107 for GRCh38 differentially expressed genes were identified using DEseq2 Assembly: CanFam3.1 genome assembly or GRCh38 human genome assembly Supplementary files format and content: tab-delimited text files include CPM normalized counts
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Submission date |
Mar 06, 2023 |
Last update date |
Dec 15, 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 |
GPL25760 |
Series (1) |
GSE226767 |
Direct Comparison of Canine and Immune Responses Using Transcriptomic and Functional Analyses |
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Relations |
BioSample |
SAMN33604297 |
SRA |
SRX19578785 |
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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