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Sample GSM7009812

Query DataSets for GSM7009812

Status

Public on Jan 30, 2023

Title

proximal_rep1

Sample type

SRA

Source name

proximal tail skin

Organism

Acomys cahirinus

Characteristics

tissue: proximal tail skin
genotype: WildType
age: newborn

Extracted molecule

total RNA

Extraction protocol

Total RNA was extracted using a RNeasy Fibrous Tissue Mini Kit (QIAGEN 74704) according to the manufacturer’s protocol.
Strand-specific TruSeqTM (Illumina) cDNA libraries were prepared from 1 µg total RNA input with a 300 bp median insert length by HudsonAlpha Discovery in Huntsville, Alabama, USA.

Library strategy

RNA-Seq

Library source

transcriptomic

Library selection

cDNA

Instrument model

Illumina NovaSeq 6000

Description

Full-thickness skin collected from the proximal end of the tail from a newborn Acomys cahirinus pup.

Data processing

Base calls and quality scores produced by the Illumina NovaSeq6000 with onboard RTA3 software.
RNA-seq FASTQ-files were quality and adapter trimmed using Trimmomatic v0.39 with the parameters : LEADING:3 TRAILING:3 SLIDINGWINDOW:4:15 MINLEN:36 and ILLUMINACLIP with the TruSeq3-PE-2.fa adapter set (available on the Trimmomatic github page: https://github.com/timflutre/trimmomatic/tree/master/adapters) and values of 2, 30,10 for seed mismatches, palindrome clip threshold and simple clip threshold, respectively.
The preprocessed reads were aligned to the Acomys skin transcriptome (GEO Accession GSE113081) with HiSat2 (v2.2.1) using default parameters for paired-end reads.
Individual read set alignments representing the output from HiSat2 were assembled into transcripts with Stringtie using default settings. Counts per transcript for each RNA-Seq sample were determined with the featureCounts function from the Rsubreads package, which mapped the aligned reads to individual transcripts within the Acomys transcriptome assembly. Featurecounts was called with the following parameters: -s 2 -p -t exon -g gene_id -T 8. Counts to isoforms that are representatives of the same trinity loci were aggregated to enable gene level expression analysis.
The Acomys transcriptome has been annotated to define Mus – Acomys orthologous for a subset of the transcripts. These annotations were used in this analysis. Differentially expressed genes were discovered with the EdgeR package (v3.36) using the aggregated count matrix as input. Count data were TMM normalized, distal/proximal tail skin samples from the same individual were treated as paired samples, and genes differentially expressed between Acomys distal and proximal tail skin samples with pvalues <= 0.05 after Benjamini & Hochberg false discovery rate correction were reported as significant.
Assembly: Trinity assembled Acomys transcriptome (GSE113081).
Supplementary files format and content: featurecounts_Acomus_Distal_proximal.csv: Comma-separated file detailing the feature counts of each Acomys sample to the Acomus Trinity assembled transcript reference (GSE113081).
Supplementary files format and content: Trinity_gene_level_osteo_SAMPLE_ReadCounts.csv: Comma-separated file describing reads mapped to the Trinity based transcriptome assembly reduced from transcript ‘isoform’ level representation to ‘gene’ level representation. Thus, counts associated with isoforms from the same gene have been aggregated to represent counts/gene.
Supplementary files format and content: Trinity_gene_level_osteo_SAMPLES_RENAMED_AllSig-ANNOTATED.csv: Comma-separated file describing EdgeR results of genes identified to be differentially expressed between DISTAL and PROXIMAL samples after Benjamini & Hochberg false discovery rate correction (pval <= 0.05).

Submission date

Jan 29, 2023

Last update date

Jan 30, 2023

Contact name

William Bradley Barbazuk

E-mail(s)

bbarbazuk@ufl.edu

Phone

352-273-8624

Organization name

University of Florida