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Sample GSM1122825

Query DataSets for GSM1122825

Status

Public on Feb 05, 2014

Title

Rat non-pineal mixed tissue, ZT7 (mid-day)

Sample type

SRA

Source name

Rat non-pineal mixed tissue, ZT7 (mid-day)

Organism

Rattus norvegicus

Characteristics

strain: Sprague-Dawley
tissue: Non-pineal mixed tissue
animals in pool: 3
time of day: ZT7 (mid-day)
age: 6-8 weeks

Treatment protocol

Rats were housed in 14:10 lighting cycle

Extracted molecule

total RNA

Extraction protocol

Total RNA was extracted with Trizol, followed by clean-up using RNeasy Micro kit with on-column DNase treatment. Quantitation and quality assuance were performed using a NanoDrop and Bioanalyzer, respectively.
A SPRI-TE kit (Beckman-Coulter) was used wherein 10 µg of total RNA was polyA-RNA-enriched using streptavidin-coated magnetic beads, followed by Covaris RNA fragmentation. Fragmented RNA was used to generate cDNA using reverse transcriptase (SuperScript II) and random primers. The cDNA was further converted into double stranded cDNA and, after an end repair process (Klenow fragment, T4 polynucleotide kinase and T4 polymerase), was ligated to Illumina paired end (PE) adaptors containing a 6-basepair bar-code. Size selection was performed using a Pippin Prep 1.5% cartridge. Libraries were enriched using 16 cycles of PCR and purified using Ampure XP Beads (Agencourt). Each library was run as one of six bar-coded samples per lane on an Illumina HiSeq machine yielding paired-end, 101-base sequencing reads. Image analysis and base calling were done using RTA1.12.4.2 and CASAVA1.8.0.

Library strategy

RNA-Seq

Library source

transcriptomic

Library selection

cDNA

Instrument model

Illumina HiSeq 2000

Description

For the mixed tissues sample, the following tissues from three Sprague-Dawley rats sacrificed at ZT7 were used: cortex, cerebellum, midbrain, hypothalamus, hindbrain, spinal cord, retina, pituitary, heart, liver, lung, kidney, skeletal muscle, small intestine, adrenal gland. Total RNA was extracted from each tissue, and then equal amounts of each of the 15 tissues were combined for the final pooled sample.

Data processing

Image analysis and base calling were done using RTA1.12.4.2 and CASAVA1.8.0.
RNA-Seq reads were aligned to the rn4 genome assembly using Tophat-1.3.1.Linux_x86_64
FPKM values were calculated using Cufflinks-1.2.0.Linux_x86_64
Genome_build: rn4
Supplementary_files_format_and_content: FPKM abundance measurements

Submission date

Apr 15, 2013

Last update date

May 15, 2019

Contact name

Steven Coon

E-mail(s)

coons@mail.nih.gov

Phone

3014516622

Organization name

NIH