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Sample GSM3273367 Query DataSets for GSM3273367
Status Public on Feb 26, 2020
Title HalfHourTreatment_2a
Sample type SRA
 
Source name Neonatal rat ventricular myocytes
Organism Rattus norvegicus albus
Characteristics agent: 10 micromol/L rosiglitazone (Sigma-Aldrich) and 1 percent dimethyl sulfoxide (DMSO)
strain: Sprague-Dawley
time point: HalfHour
Treatment protocol The control group was supplemented with 1% dimethyl sulfoxide (DMSO) and the experimental group was supplemented with 10 μmol/L rosiglitazone (Sigma-Aldrich) and 1% dimethyl sulfoxide (DMSO). NRVMs were exposed to control and experimental conditions for 0.5 hr, 24 hr, and 48 hr.
Growth protocol All animal procedures were in accordance with the San Diego State University Animal Subjects Committee (UASC) and NIH Animal Welfare Assurance A3728-01. Neonatal rat ventricular myocytes (NRVMs) were harvested from 1-day old Harlan Sprague-Dawley Rats (Rattus norvegicus albinus) and cultured as previously described by Sprenkle et al 1995. Preplating isolated cells to remove fibroblasts yielded >98% NRVMs. Cell count was determined after preplating. Pooled NRVMs were centrifuged and resuspended in 20 ml DMEM/F-12 (Gibco, Life Technologies, CA, USA) with 10% fetal bovine serum (Irvine Scientific, CA, USA) and quantified via hemacytometer. NRVMs were plated on Fibronectin (Gibco)- coated 35 mm dishes and incubated overnight to allow for recovery. On the following day, NRVMs were washed twice with 1:1 medium DMEM/F-12, kanamycin, ampicillin, and fungizone. NRVMs were incubated overnight in minimal medium consisting of 1:1 medium with 1 mg/ml bovine serum albumin (Sigma, MO, USA). On the third day, cells were washed two times with 1:1 medium. Minimal medium was replaced with Insulin-Transferrin-Selenium medium consisting of minimal medium, 1X Insulin-Transferrin-Selenium (Life Technologies), 0.4X MEM nonessential amino acids mixture (Sigma), and 0.1X MEM vitamins medium (Invitrogen, CA, USA).
Extracted molecule total RNA
Extraction protocol After the appropriate exposure time, isolation of total RNA for RNA sequencing from
NRVMs was performed using RNeasy Mini Kit (Qiagen) as per the manufacturer’s protocol. RNA purity was determined by spectrophotometry using a NanoDrop 2000 (Thermo Scientific).
Library were prepared using the NEBNext® Ultra II™ Directional RNA Library Prep Kit for Illumina (New England Biolabs Inc) and procedures were made are described by the manufacturer.
 
Library strategy RNA-Seq
Library source transcriptomic
Library selection cDNA
Instrument model Illumina NextSeq 500
 
Description 03_HalfHourTreatment_2a
Treatment
Data processing Quality check of the Fastq files using FastQC v0.11.5
Sequenced reads (fastq files) were trimmed for adaptor sequence, nucleotides with the phred score below 30 and for overrepresented sequences using cutadapt, version 1.11
The resulting SAM files were sorted and inputted into the Python package HTSeq to generate count data for gene-level differential expression analyses.
Transcript count data from DESeq2 analysis of the samples were sorted according to their adjusted p-value or q-value, which is the smallest false discovery rate (FDR) at which a transcript is called significant.
The analysis was carried out on an OnRamp Bioinformatics Genomics Research Platform (OnRamp Bioinformatics, San Diego, CA). OnRamp’s advanced Genomics Analysis Engine utilized an automated RNAseq workflow to process the data, including data validation and quality control and read alignment to the human genome (hg19).
genome build: Rnor_6.0
processed data files format and content: The processed data files are “.txt” tabular files, contain in as column the identifiers ensembl_gene_id, external_gene_name, description, hsapiens_homolog_ensembl_gene, hsapiens_homolog_perc_id, hgnc_symbol, Human_description, Human_entrez_geneid, ensembl_peptide_ids, ensembl_transcript_ids, with annotation for ENSEMBLE and gene symbols, the original murine gene and its orthologous genes in humans. In the sequences we have columns with the sample names and the raw counts of the genes mentioned. The last section of the files possess 4 columns: baseMean log2FoldChange, lfcSE ,stat, pvalue, padj. Showing the results from DESeq2 analysis.
 
Submission date Jul 17, 2018
Last update date Mar 26, 2021
Contact name Gary Hardiman
E-mail(s) hardiman@musc.edu, g.hardiman@qub.ac.uk, glen@musc.edu
Organization name Medical University of South Carolina
Department Medicine
Street address 135 Cannon Street
City Charleston
State/province SC
ZIP/Postal code SC
Country USA
 
Platform ID GPL25338
Series (1)
GSE117198 A Time-series transcriptome analysis of Rosiglitazone effects in mice neonatal cardiomyocytes.
Relations
BioSample SAMN09770049
SRA SRX4513900

Supplementary data files not provided
SRA Run SelectorHelp
Raw data are available in SRA
Processed data are available on Series record

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