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Sample GSM1861766

Query DataSets for GSM1861766

Status

Public on Aug 27, 2015

Title

Ishikari Bay, male 2, OE I5

Sample type

SRA

Source name

Ishikari Bay_male_olfactory rosette

Organism

Oncorhynchus keta

Characteristics

origin of specimen: caught on Oct. 2, 2011 at the coastal zone of the Pacific Ocean in the Ishikari Bay (43°23’ N, 141°13’ E)
gender: male
tissue: olfactory rosette

Extracted molecule

total RNA

Extraction protocol

Olfactory rosettes were obtained from each individual, flash frozen in liquid nitrogen and stored at -80 °C till RNA extraction. Total RNA was isolated using TRIzol (Invitrogen, Carlsbad, USA) and analyzed with an Agilent Bioanalyzer 2100 total RNA Nano series II chip (Agilent). A transcriptome library for each individual was prepared from 2 microgram total RNA, using the Illumina TruSeq™ RNA Sample Prep Kit v2 following the manufacturer’s instructions (Illumina Inc.).
RNA libraries were prepared for sequencing using standard Illumina protocols

Library strategy

RNA-Seq

Library source

transcriptomic

Library selection

cDNA

Instrument model

Illumina HiSeq 2500

Description

processed data files:
DiffExpr_Location_7-12_vs_1-6_Salmo_salar_3_mismatches.xlsx;
DiffExpr_Location_7-12_vs_1-6_Oncorhynchus_keta_de_novo.xlsx

Data processing

Illumina Casava software used for basecalling: SCS2.2.38/RTA1.18.61
In strategy 1, reads were aligned to 48,223 Salmo salar cDNA sequences (downloaded from NCBI) allowing 3 mismatches per 50 nt read (3 mismatches ~ 94% identity) using TopHat (version 2.0.5) (Trapnell et al., 2009)
In strategy 2, reads were aligned to a de novo assembled Chum salmon olfactory rosette contig reference file using TopHat (version 2.0.5) (Trapnell et al., 2009)
The resulting files were filtered using SAMtools (version 0.1.18) (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 (Anders et al., 2014)
Differentially expressed genes were identified using DESeq (Anders and Huber, 2010) and data were further processed using Microsoft Excel

DiffExpr_Location_7-12_vs_1-6_Oncorhynchus_keta_de_novo.xlsx;
The reference FASTA file (Oncorhynchus_keta_cDNA.fa) was made as follows: A total of ~250 million paired-end 2 x 50 nt reads (~25 Gb data) from all 12 RNAseq libraries were combined and used in a single de novo assembly to generate cDNA sequences corresponding to mRNAs that are expressed in the O.keta olfactory placodes. This resulted in 98,542 cDNA contigs ranging in size from 200 - 17,327 nt.

DiffExpr_Location_7-12_vs_1-6_Salmo_salar_3_mismatches.xlsx:
The reference FASTA file consisted of 48,223 Salmo salar cDNA sequences downloaded from NCBI, and thus the identifiers in the *mismatches.xlsx (GI numbers) are searchable in http://www.ncbi.nlm.nih.gov/nuccore
Supplementary_files_format_and_content: Microsoft Office Excel file (.xlsx) with the following columns: id; Function description; sample 7; sample 8; sample 9; sample 10; sample 11; sample 12; sample 1; sample 2; sample 3; sample 4; sample 5; sample 6; BaseMean; baseMeanA; baseMeanB; foldChange; log2FoldChange; pval; padj

Submission date

Aug 26, 2015

Last update date

May 15, 2019

Contact name

Ron P Dirks

E-mail(s)

dirks@futuregenomics.tech

Phone

+31-71-2074610

Organization name

Future Genomics Technologies BV