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Sample GSM3025460

Query DataSets for GSM3025460

Status

Public on Apr 04, 2019

Title

BRN_CFA_rep1

Sample type

SRA

Source name

brain

Organism

Mus musculus

Characteristics

strain: BALB/c
tissue: brain
age: 18 weeks
pain model: CFA

Treatment protocol

Complete Freund’s adjuvant (CFA): (CFA; 50%; Sigma) was injected subcutaneously in a volume of 20 μl into the left and right plantar hind paws using a 100-μl microsyringe with a 30-gauge needle. 3 days post-CFA mice were sacrificed. Spared nerve injury (SNI): performed under isoflurane/oxygen anesthesia using an operating microscope (x40), the three terminal branches of the sciatic nerve (tibial, sural and common peroneal) are exposed. The tibial and common peroneal nerve are cut, after tight ligation with 6.0 silk, “sparing” the sural nerve. The incisions are closed in layers using interrupted sutures (6-0 Vicryl) and wound clips. The animal recovers on a thermostatically-controlled heating pad (carefully monitored to prevent over-heating) until ambulatory as per standard operating procedures. To reduce the overall number of mice required, the SNI surgery was performed in a bilaterally fashion (i.e. left and right side). 7 days after the surgery the mice were sacrificed.

Extracted molecule

polyA RNA

Extraction protocol

Total RNA was isolated using RNeasy Lipid Tissue Mini Kit, while RNA from blood cells was isolated using the RNeasy Mini Kit, including DNase I treatment (all Qiagen), all according to the manufacturer’s instructions. Total RNA was quantified using the NanoDrop 2000 (Thermo Scientific), while RNA quality was assessed with the 2100 Bioanalyzer (Agilent Technologies).
All RNA-sequencing procedures were performed by Genomics Platform facility at Institute for Research in Immunology and Cancer, Montreal, Canada. Transcriptome libraries were generated from 1 μg of total RNA using the Kapa RNA stranded Sample Prep Kit (KK8400, KAPABiosystems) following the manufacturer’s protocols. Briefly, poly-A mRNA was purified using poly-T oligo-attached magnetic beads using two rounds of purification. During the second elution of the poly-A RNA, the RNA was fragmented and primed for cDNA synthesis. During cDNA synthesis, dUTP is incorporated in the second-strand synthesis, where dUTP-containing strand is selectively degraded. Adenylation of the 3’ ends and ligation of adapters were done following the manufacturer protocol. Enrichment of DNA fragments that have adapter molecules on both ends was done using 10 cycles of PCR amplification using the KAPA PCR mix and Illumina-adapted primers cocktail.

Library strategy

RNA-Seq

Library source

transcriptomic

Library selection

cDNA

Instrument model

Illumina HiSeq 2000

Description

Parisien_Suppl_Table_01_gene_BRN_CTRvsCFA.xlsx

Data processing

RNA-Seq data has been trimmed with Trimmomatic version v0.32.
Trimmed reads were aligned using tophat version v2.0.11 and bowtie version v1.0.0 on the reference mouse genome version mm10.
Differential gene expression was estimated with cuffdiff version v2.2.1.
Genome_build: mm10
Supplementary_files_format_and_content: tab-delimited text files include fpkM values, fold changes, and associated fold change P-values, for paired CFA versus CTR, or SNI versus CTR.

Submission date

Feb 27, 2018

Last update date

Apr 04, 2019

Contact name

Marc Parisien

E-mail(s)

marc.parisien@mcgill.ca

Organization name

McGill University

Department

Dentistry