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Sample GSM3258484

Query DataSets for GSM3258484

Status

Public on Jul 06, 2018

Title

19G-CTRL rep 2

Sample type

SRA

Source name

Testis

Organism

Oreochromis niloticus

Characteristics

tissue: Testis
age: One year old
Sex: male

Treatment protocol

Tilapia were randomly assigned to the BaP-treated or control groups (n = 24 each) and kept distributed in four experimental aquariums (2 for control group and 2 for BaP-treated with 12 fish per aquarium). Male adult tilapia received 5 separate i.p injections (a single injection every 6 days) of BaP (3 mg/kg) or DMSO + corn oil as the carrier solution. Two days after the 5th injection (Day 26), six fish from each BaP-treated and control groups were euthanized using an overdose of 0.5 g/l MS-222 (Sigma Aldrich, Inc). Liver and testis were excised, weighed, snap frozen in liquid nitrogen and then stored at -80 °C for subsequent molecular analyses.

Growth protocol

Adult male tilapias (378.8 ± 9.63 g in weight and 27.7 ± 0.214 cm in length) were selected from one year old tilapia fish stocks maintained at CINVESTAV-Merida. After selection, the fish were kept for a four-week acclimation period in a 300 l aquarium at ~ 28°C with aerated water, natural photoperiod and flow-through conditions.

Extracted molecule

total RNA

Extraction protocol

Total RNA was isolated from ~0.5 mg of liver or testis tissues using the Trizol reagent method according to the manufacturer's procedure (Invitrogen™, Carlsbad CA,USA). The RNA pellet was resuspended in 20–50 μl of RNAsecureTM Reagent (Ambion Austin, TX, USA) and DNAse was-treated with Turbo DNA-free (Ambion Austin, TX, USA). RNA cleanup was performed using Phenol-chloroform (1:1) extraction, followed by sodium acetate (3M) and ethanol (100 %) precipitation (Chomczynski et al., 1987).
RNA libraries were prepared for sequencing using standard Illumina protocols. Samples were processed following the manufacturer’s protocol for NEBNext Ultra™ RNA Library Prep Kit for Illumina (New England, Biolabs (NEB), lpswich, MA, USA) in conjunction with the NEBNext Poly(A) mRNA Magnetic Isolation Module and the NEBNext Multiplex Oligos for Illumina (Index Primers Set 1)

Library strategy

RNA-Seq

Library source

transcriptomic

Library selection

cDNA

Instrument model

Illumina NextSeq 500

Description

MTG.txt
Sample8_Ind16-35365336

Data processing

Tilapia RNA-Seq reads were trimmed (adaptor sequences) and reads of low quality (< 12 bp) were eliminated. The reference genome from O. niloticus (Orenil2.0) were downloaded from the Ensembl web site (http://www.ensembl.org/Oreochromis_niloticus/Info/Index). Clean reads were aligned to the reference genome using Tophat and the resulting Binary Sequence Alignment/Map (BAM) files were quantified for gene/transcript expression. Differential expression (DE) analysis of the clean reads was performed by testing the BaP treated read counts against controls read counts using exact test with R package EdgeR with a significance threshold of p < 0.05; fold change greater than ± 1. Pathway analysis was performed using PathwayStudioTM V9 (Elsevier, Inc., Rockville, MD, USA) operating with the ResNet 10.0 database. The gene set enrichment analysis (GSEA) and sub-network enrichment analysis (SNEA) using the Mann-Whitney test with an alpha level of p < 0.05 were used to identified potential pathways altered by BaP exposure.
Genome_build: Orenil2.0

Submission date

Jul 05, 2018

Last update date

Jul 06, 2018

Contact name

Reyna Cristina Colli-Dula

E-mail(s)

[email protected]

Organization name

CINVESTAV

Department

Recursos del Mar