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GEO help: Mouse over screen elements for information. |
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| Status |
Public on Sep 13, 2017 |
| Title |
RNA-seq for isolated mouse brain microglia, LPS group, replicate 3 |
| Sample type |
SRA |
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| Source name |
isolated brain microglia
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| Organism |
Mus musculus |
| Characteristics |
sample type: replicate 3
strain: C57BL/6
genotype/variation: WT
agent: LPS
tissue: brain
cell type: microglia
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| Treatment protocol |
To activate microglia, adult wildtype B6 mice received daily intraperitoneally injection of 1mg/Kg lipopolysaccharide (L4524, Sigma) or phosphate-buffered saline for 4 consecutive days before sacrifice.
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| Extracted molecule |
total RNA |
| Extraction protocol |
For Ptbp2-KO brain cortex RNA-seq, E18.5 mouse cortex RNA (three biological replicates) from Ptbp2-KO mice (Licatalosi et al., 2012) was prepared using Trizol (Ambion); ribosomal RNA was removed from 1 μg RNA using Ribo-Zero rRNA removal Kit (Illumina). For microglia isolation, mice were perfused with 30ml of heparinized PBS. Brains were excised, minced and two brains were placed into C tube and processed as per instructions for the Neural Tissue Dissociation Kit P (Miltenyi Biotec) using the gentleMACS Octo Dissociator with Heaters (Miltenyi Biotec, protocol 37_ABDK). Once digested, the debris and myelin was removed using the Debris Removal Solution (Miltenyi Biotec) followed by CD11b positive selection using Microglia CD11b beads (Miltenyi). Cells were counted, washed in PBS and pelleted prior to isolation of RNA. Total RNA from isolated microglia was prepared using Trizol (Ambion) and High Pure RNA Isolation Kit (Roche). The Total RNA was then further purified for polyadenylated RNA by using 150ng Total RNA in Dynabeads mRNA Purification Kit (Ambion).
RNA-seq libraries were prepared using TruSeq RNA Sample Preparation Kit v2 (Illumina) following manufacturer’s instructions.
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina MiSeq |
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| Data processing |
RNA-seq mapping to mouse genome (mm10) was performed using STAR aligner (Dobin et al., 2013).
Genome_build: mm10
Supplementary_files_format_and_content: txt files for read counts at gene level (Ensembl) generated by STAR aligner
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| Submission date |
Jun 26, 2017 |
| Last update date |
May 15, 2019 |
| Contact name |
Hun-Way Hwang |
| E-mail(s) |
Hunway.Hwang@pitt.edu
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| Organization name |
University of Pittsburgh
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| Department |
Department of Pathology
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| Lab |
S754 Scaife Hall
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| Street address |
3550 Terrace Street
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| City |
Pittsburgh |
| State/province |
PA |
| ZIP/Postal code |
15261 |
| Country |
USA |
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| Platform ID |
GPL16417 |
| Series (1) |
| GSE94054 |
cTag-PAPERCLIP Reveals Alternative Polyadenylation Promotes Cell-Type Specific Protein Diversity and Shifts Araf Isoforms with Microglia Activation |
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| Relations |
| BioSample |
SAMN07276964 |
| SRA |
SRX2955626 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM2683278_M7.LPS3.ReadsPerGene.out.tab.c1c2.txt.gz |
183.8 Kb |
(ftp)(http) |
TXT |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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