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Sample GSM7098249

Query DataSets for GSM7098249

Status

Public on Dec 27, 2023

Title

ID16363_liver_0yr

Sample type

SRA

Source name

liver

Organism

Papio anubis

Characteristics

tissue: liver
animal id: 16363
treatment: LCLF
time: 0yr

Treatment protocol

The animals were feeding a high cholesterol, high fat diet for two years, switching from a low cholesterol, low fat diet.

Extracted molecule

total RNA

Extraction protocol

Approximately 10 mg of each frozen tissue sample was homogenized in 1ml QIAzol Lysis Reagent (79306, QIAGEN) using BeadBug 6 Microtube Homogenizer (D1036, Benchmark Scientific) with 1.5mm Zirconium beads (D1032-15, Benchmark Scientific). RNA was immediately extracted from the homogenate using QIAGEN RNeasy Universal Kit (73404, QIAGEN). RNA concentration and quality was measured using Agilent RNA 6000 Nano Kit and RNA 6000 Pico Kit (5067-1511 & 5067-1513, Agilent) on Agilent 2100 Bioanalyzer.
TruSeq RNA Library Preparation Kit v2 (RS-122-2001, RS-122-2002, Illumina); TruSeq stranded mRNA Library Preparation Kit (20020595, Illumina)
96 libraries from 16 animals in batch 1 were performed using TruSeq RNA Library Preparation Kit v2 (RS-122-2001, RS-122-2002, Illumina) and were sequenced 50 base pairs, single-end using the Illumina HiSeq4000 according to manufacturer instructions with the goal of achieving at least 15 million reads per sample. Each sequencing lane contained 24 multiplexed samples from different RNA extraction batches. 498 libraries from 83 animals in batch 2 were performed using TruSeq stranded mRNA Library Preparation Kit (20020595, Illumina) and were sequenced 50 base pairs, paired-end using the Illumina NovaSeq6000 according to manufacturer instructions with the goal of achieving at least 30 million reads per sample. Each sequencing lane contained 84 multiplexed samples from different RNA extraction batches. Samples from the same animal were always in the same RNA extraction, library preparation, and sequencing batches. Samples from different animals were randomized in every processing step to avoid potential batch effects.

Library strategy

RNA-Seq

Library source

transcriptomic

Library selection

cDNA

Instrument model

Illumina HiSeq 4000

Data processing

RNA-seq data were aligned to the baboon reference genome Panubis1.0 with STAR v2.7.7a. FASTQC was used to confirm that the reads were of high quality. Gene-level expression quantification was performed using featureCounts. Quality controls were performed on two levels using a similar pipeline for GTEx. On the sample level, we first identified and removed sample swaps and contaminations where a sample was not matched to its genotype or contained a mixture of two or more samples using verifyBamID. We removed outlier samples based on expression profile using PCA. We checked sex correctness by examining gene expression for genes on the Y chromosome for each sample. On the gene level, gene expression values for all samples from a given tissue were normalized using the following procedure: 1) read counts were normalized between samples using trimmed mean of M-values (TMM); 2) genes were selected based on expression thresholds of ≥0.1 TPM in ≥10% of samples, and > 3 reads (unnormalized) in ≥10% of samples for both diet conditions; 3) expression values for each gene were inverse normal transformed across samples. After the quality controls, we obtained 570 high-quality RNA samples and 20,224 genes for downstream analyses.
Assembly: Panubis1.0
Supplementary files format and content: raw count matrix of all samples (n=588)
Supplementary files format and content: raw count matrix of post-QC samples (n=570)

Submission date

Mar 14, 2023

Last update date

Dec 27, 2023

Contact name

Wenhe Lin

E-mail(s)

wenhelin@uchicago.edu

Organization name

University of Chicago

Street address

920 E 58th St