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Sample GSM5241654

Query DataSets for GSM5241654

Status

Public on Nov 24, 2021

Title

FG1907_03

Sample type

SRA

Source name

rete mirabile tissue

Organism

Anguilla anguilla

Characteristics

developmental stage: silver eel
tissue: rete mirabile

Treatment protocol

Eels were anesthetized with 2-phenoxyethanol (1 ml*L^-1), and subsequently decerebrated and spinally pithed. The body wall was opened ventrally, the swimbladder was exposed and carefully freed from connective tissue. Artery and vein at the heart pole of the retia mirabilia were carefully separated and occlusively cannulated using PE50 and PE60 catheters, respectively. Rete vessels were perfused with heparinized (100 i.u*mL^-1) saline solution containing (in mmol*L^-1) NaCl, 129; KC1, 5; MgSO4, 0.9; CaCl2, 1.1; glucose, 15. Perfusion of rete vessels was controlled by continuous binocular inspection and both retia mirabilia were dissected immediately after total clearance of blood cells. Half of the tissue was blotted dry on absorbent paper, shock frozen in liquid nitrogen and stored at -80°C until further use for analysis of protein expression. The second half was transferred into 1 ml RNAlater™ solution and immediately shock frozen in liquid nitrogen for analysis of the transcriptome.

Growth protocol

European eels were caught in Lake Constance by local fishermen and kept in freshwater for a few days.

Extracted molecule

total RNA

Extraction protocol

Total RNA was isolated from gas gland tissue using the Qiagen miRNeasy kit. Quality and integrity of the RNA waere checked on an Agilent Bioanalyzer 2100 total RNA Nano series II chip.
Illumina RNASeq libraries were prepared from 0.5 µg total RNA using the Illumina TruSeq Stranded mRNA Library Prep.

Library strategy

RNA-Seq

Library source

transcriptomic

Library selection

cDNA

Instrument model

Illumina NovaSeq 6000

Data processing

Illumina NovaSeq6000 RTA3 software used for basecalling: version 3.4.4
For comparison between the experimental groups, all forward reads were trimmed to a final size of 50 nt using fastx_trimmer (http://hannonlab.cshl.edu/fastx_toolkit/index.html)
Reads were aligned to the draft genome sequence of the European eel (Henkel et al., 2012) FASTA with transcript sequences is available on the series record. using TopHat (version 2.0.13) (Trapnell et al., 2009)
The resulting files were filtered using SAMtools (version 1.2) (Li et al., 2009) to exclude secondary alignment of reads
Aligned fragments per predicted gene were counted from SAM alignment files using the Python package HTSeq (version 0.6.1p1) (Anders et al., 2015)
As a correction scaling factor, library size estimates, determined using the R/Bioconductor (release3.3.2) package DESeq (Anders and Huber, 2010), were employed.
Genome_build: Anguilla_anguilla_genome_v1_09_nov_10 (GCA_000695075.1) (Henkel et al., 2012) FASTA with transcript sequences is available on the series record.
Supplementary_files_format_and_content: Microsoft Office Excel file (.xlsx) with the following columns: id; Name; Best hit; Description; Alternate description; Min e-value; Similarity; GO molecular function; GO biological process; GO cellular component; baseMean; baseMeanA; baseMeanB; foldChange; log2FoldChange; pval; padj; sample x; sample y; sample z

Submission date

Apr 14, 2021

Last update date

Nov 24, 2021

Contact name

Gabriel Schneebauer

Organization name

University of Innsbruck

Street address

Technikerstrasse 25

City

Innsbruck