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| Status |
Public on Sep 18, 2017 |
| Title |
Experimental Rep3 |
| Sample type |
SRA |
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| Source name |
Whole Ear
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| Organism |
Mus musculus |
| Characteristics |
strain: C57Bl/6
tissue: Skin
genotype: NaV1.8+ Jak1F/F
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| Extracted molecule |
total RNA |
| Extraction protocol |
Samples of murine skin were obtained and stored in RNAlater (Sigma) before processing following the manufacturer’s instructions. To extract whole tissue RNA, samples were homogenized using a bead homogenizer and processed using the Qiagen RNeasy kit following the manufacturer’s instructions. Following total RNA extraction, samples were treated with DNase (Turbo DNA-Free Kit, Thermo Scientific) following the manufacturer’s instructions before sequencing library preparation.
Library preparation was performed with 1 ug of total RNA, integrity was determined using an Agilent Bioanalyzer. Ribosomal RNA was removed by a hybridization method using Ribo-ZERO kits (Illumina-EpiCentre). mRNA was then fragmented in buffer containing 40 mM Tris Acetate (pH 8.2), 100 mM Potassium Acetate, and 30mM Magnesium Acetate and heated to 94°C for 150 seconds. mRNA was reverse transcribed to yield cDNA using SuperScript III RT enzyme (Life Technologies, per manufacturer’s instructions) and random hexamers. A second strand reaction was performed to yield ds-cDNA. cDNA was blunt ended, had an A base added to the 3’ ends, and then had Illumina sequencing adapters ligated to the ends. Ligated fragments were then amplified for 12 cycles using primers incorporating unique index tags. Fragments were sequenced on an Illumina HiSeq-3000 using single reads extending 50 bases.
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 3000 |
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| Data processing |
RNA-sequencing reads were aligned to the Ensembl release 76 assembly with STAR version 2.0.4b. Gene counts were derived from the number of uniquely aligned unambiguous reads by Subread:featureCount version 1.4.5. Transcript counts were produced by Sailfish version 0.6.3. Sequencing performance was assessed for total number of aligned reads, total number of uniquely aligned reads, genes and transcripts detected, ribosomal fraction known junction saturation and read distribution over known gene models with RSeQC version 2.3.
Genome_build: mm10
Supplementary_files_format_and_content: Tab-delimited text files including count values for each sample.
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| Submission date |
Jun 22, 2017 |
| Last update date |
May 15, 2019 |
| Contact name |
Landon Kyle Oetjen |
| E-mail(s) |
lkoetjen@wustl.edu
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| Organization name |
Washington University
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| Street address |
660 S Euclid Avd
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| City |
St. Louis |
| State/province |
MO |
| ZIP/Postal code |
63110 |
| Country |
USA |
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| Platform ID |
GPL21493 |
| Series (1) |
| GSE100397 |
Sensory neurons co-opt classical immune signaling pathways to mediate chronic itch [6 NaV1.8+ Il4raF/F mice treated with MC903 compared to skin from 4 littermates] |
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| Relations |
| BioSample |
SAMN07270012 |
| SRA |
SRX2948708 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM2680303_gene_counts_AAACCTT.txt.gz |
2.7 Mb |
(ftp)(http) |
TXT |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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