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Sample GSM2629045

Query DataSets for GSM2629045

Status

Public on Feb 08, 2018

Title

PRA_ZT08

Sample type

SRA

Source name

Preoptic area

Organism

Papio anubis

Characteristics

tissue: Preoptic area
time: ZT08

Extracted molecule

polyA RNA

Extraction protocol

Twelve male Papio anubis hamadryas (age 5-6 yrs, 7-11 kg) were maintained in a 12hr light-12 hr dark cycle. Animals were fed twice a day (at 9:00 and 15:00) with fruits and primate meal pellets, with access to water ad libitum. Timed biosamples were collected every 2 hrs from ZT0 to ZT22, one primate at each sample time. The Institute veterinary services administered animals a lethal dose of sodium pentobarbital, after which blood (centrifuged for plasma), urine and CSF samples collected. Animals were perfused through the left ventricle with 4 liters of ice cold (2°C) buffered, oxygenated Ame's solution. Collection of brain sections used standardized procedures for NHPs (Davenport et al., 2013). The brain and eyes were rapidly removed (less than 5 min) and kept at 4°C on ice. Tissue collections were made simultaneously in three stations. In Station 1, the brain was cut into serial sections using a specifically built baboon brain matrix. Sections were placed on individual ice cold plates, each section photographed and punches obtained from selected brain structures. Sections were packaged in individual sterilized plastic pouches (Nasco) then frozen in dry ice and stored at -80°C. At the same time (Station 2), samples were taken from the eyes (retina, RPE, vitreous, lens, cornea, iris, optic nerve), divided into aliquots and frozen in dry ice. During the night-time periods, the head and eyes were covered with opaque plastic prior to perfusion and dissections of the eye and retina were performed in the dark using infrared viewers. Station 3 collected various body tissues and organs. All tissues were distributed into individual aliquots with certain larger tissues stored in bulk samples (liver, muscle, etc.), frozen in dry ice, and then stored at -80°C. The whole procedure, from collection to storage lasted no more than 30-40 min.
Libraries were prepped using Illumina’s TruSeq Stranded mRNA HT kit according to manufacturer’s instructions. In brief, total RNA starting with 1ug was poly-A selected, fragmented by metal-ion hydrolysis and then converted to cDNA using SuperScript II. The cDNA was then end-repaired, adenylated and ligated with Illumina sequencing adapters. Finally, the libraries were enriched by PCR amplification. Libraries were pooled and sequenced using an Illumina HiSeq 2500 with 50-bp single-read chemistry.

Library strategy

RNA-Seq

Library source

transcriptomic

Library selection

cDNA

Instrument model

Illumina HiSeq 2500

Data processing

Raw sequencing reads were aligned to the baboon PapAnu2.0 genome with STAR
Read counts per gene were calculated using FeatureCounts
DESeq2 was used to generate FPKM values
Genome_build: PapAnu2.0
Supplementary_files_format_and_content: comma-separated values containing FPKM quantification for all samples
Supplementary_files_format_and_content: gene-level quantification (FPKM normalization)

Submission date

May 16, 2017

Last update date

May 15, 2019

Contact name

Max Chang

E-mail(s)

[email protected]

Organization name

University of California, San Diego

Street address

9500 Gilman Drive