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Sample GSM7635199

Query DataSets for GSM7635199

Status

Public on Jul 04, 2024

Title

uninfected perch spleen 43

Sample type

SRA

Source name

spleen

Organism

Perca fluviatilis

Characteristics

tissue: spleen
Sex: Female
infection status: uninfected

Extracted molecule

total RNA

Extraction protocol

Tissue samples were crushed with a steel mortar and pestle to produce a homogenized powder, while frozen spleen tissue was mechanically crushed (the tissue consistency of spleen samples was not conducive to mortar and pestle homogenization) using a Retsch Mixer Mill MM 400 (Retsch). Total RNA was extracted using a NucleoSpin RNA extraction kit (MACHEREY-NAGEL, Duren, Germany). RNA sample concentrations were measured with a NanoDrop 2000 (ThermoFisher), and sample quality was evaluated using a TapeStation 2200 (Agilent Technologies)
Sequencing libraries were prepared at the Novogene Cambridge Science Park (Cambridge, United Kingdom) from 300 ng total RNA

Library strategy

RNA-Seq

Library source

transcriptomic

Library selection

cDNA

Instrument model

Illumina NovaSeq 6000

Data processing

The raw sequence files in fastq format returned from Novogene were run through FastQC v0.11.9 both before and after trimming to check the overall quality of the reads. The trimming was performed by fastp v.0.20 . Only reads which were both longer than 50 bp and were of mean quality of 25 were processed further.
The reads which passed the quality control were mapped on the reference genome of Perca fluviatilis using hisat2 v.2.1.1 using default parameters. The produced .sam files were subsequently processed with samtools v.1.10. Lastly, the mapped reads were extracted in bam format for subsequent differential expression analysis.
Analysis for differentially expressed genes (DEGs) was performed in R v.4.2.2 using the DESeq2 v.1.36.0 package. When designing the DESeq dataset object, we considered infection status and population as fixed factors, and the DESeq model internally corrects for potential differences in library sizes. Genes with an adjusted p-value (p-adj) ≤ 0.05 [40] were considered as significantly differentially expressed between infected and uninfected individuals. Human orthologue genes were obtained via Ozerov et al., and DEGs without human orthologues were manually searched using BLAST.
Supplementary files format and content: processed fastq files

Submission date

Jul 18, 2023

Last update date

Jul 04, 2024

Contact name

Anti Vasemägi

Organization name

Swedish University of Agricultural Sciences

Street address

Stångholmsvägen 2

City

Stockholm