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Sample GSM1669536 Query DataSets for GSM1669536
Status Public on Jul 15, 2015
Title repo_micro_rep3
Sample type SRA
 
Source name Repopulating microglia
Organism Mus musculus
Characteristics tissue: Repopulating microglia
genotype: Repopulating microglia
strain: DTRmg
Treatment protocol Tamoxifen (TAM; Sigma) was suspended on 37°C in olive oil for 60 mins (MP Biomedicals). 2 mg Tamoxifen was administered subcutaneously (s.c.) twice on postnatal days 12 and 14. For microglia depletion, mice were injected intraperitoneally (i.p.) with 500ng diphtheria toxin (DT; Merck Millipore) three times, with a one-day interval between each injection.
Extracted molecule total RNA
Extraction protocol Mice were anesthetized and perfused with NaCl and organs were excised out. Brains were either fixed with 4% of PFA for histology, snap frozen in liquid nitrogen for RNA preparation, or digested according to manufacturer’s guidelines supplied in the MACS tissue dissociation kit Papain (Miltenyi Biotec). The tissue homogenate was loaded on a 30:70% Percoll gradient for enrichment of CNS infiltrates. The interphasic cellular ring was collected for further investigation. The different microglia samples were FACS sorted (FACS Aria) and immediately centrifuged down for 15 mins and 400 x g. RNA was isolated with the RNeasy® Plus Micro Kit (Qiagen) according to the manufacturer’s instructions. RNA amounts were measured by Q-bit 2.0 fluorometer (Life Technologies) and the quality was assessed on a Bioanalyzer (Agilent) using a High Sensitivity Chip. Samples with a RNA integrity number (RIN) > 8 were used for library preparation with TruSeq® RNA Sample Prep v2 Kit (Illumina) according to the manufacturer’s manual. Library size distribution was assessed on a Bioanalyzer 2100 using a High Sensitivity DNA Chip (Agilent).
RNA libraries were prepared for sequencing using standard Illumina protocols
 
Library strategy RNA-Seq
Library source transcriptomic
Library selection cDNA
Instrument model Illumina MiSeq
 
Data processing Sequenced reads were mapped to the mouse genome reference sequence (mm10, retrieved from Illumina iGenomes); TopHat version 2.0.10, combined with Bowtie2 (2.1.0.0) was used for this purpose. Default parameters were used to perform the alignment.
Quantification of gene expression was performed with the htseq-count tool from the HTSeq software suite. The corresponding annotation was also retrieved from the Illumina iGenomes archive.
Differential expression analysis was performed by using DESeq (version 1.14.0). A False Discovery Rate (FDR) cutoff of 0.05 was selected.
Genome_build: mm10
Supplementary_files_format_and_content: Tag-delimited text file, containing normalized read counts, stored in a NxM matrix (N genes, M samples) with a header line
 
Submission date Apr 28, 2015
Last update date May 15, 2019
Contact name Ari Waisman
E-mail(s) waisman@uni-mainz.de
Organization name Institute for Molecular Medicine
Street address Obere Zahlbacher Straße 67
City Mainz
ZIP/Postal code 55131
Country Germany
 
Platform ID GPL16417
Series (1)
GSE68376 Interleukin-1 signaling is involved in local self-renewal and maintenance of microglia
Relations
BioSample SAMN03571211
SRA SRX1013028

Supplementary data files not provided
SRA Run SelectorHelp
Raw data are available in SRA
Processed data are available on Series record

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