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Sample GSM5277389 Query DataSets for GSM5277389
Status Public on Sep 08, 2021
Title CRE_ZT10_r2
Sample type SRA
 
Source name Liver
Organism Mus musculus
Characteristics strain: Nr3c1fl/fl
aav injection: AAV-tbg-CRE
treatment: unfed
circadian timepoint: ZT10
tissue/cell type: Liver
molecule subtype: rRNA depleted RNA
Treatment protocol Nr3c1fl/fl mice were injected with AAV-tbg-GFP or AAV-tbg-CRE ten days before sacrifice. The primary hepatocytes were serum depleted in Gibo Medium 199 + GlutaMAX supplemented with 1 % P/S and 0.1 % HAS for 4 hours before cells were stimulated with 500 nM dex for 2.5 hours, human insulin for 2 hours or in a combination and RNA was harvested.
Growth protocol Mice were housed at 12 hour light/dark cycles and trained to night restricted feeding. Primary hepatocytes were isolated from Male Sprague Dawley rats and seeded on collagen coated plates in Gibo Medium 199 + GlutaMAX (Gibco life technologies, cat: 41150-020) supplemented with 100 nM. Decadron (dex), 1 % P/S, 4 % FCS and 1 nM human insulin. 2-3 hours after isolation, the medium was changed to Gibo Medium 199 + GlutaMAX supplemented with 1 % P/S, 0.1 % FCS, 1 nM human Insulin and kept overnight.
Extracted molecule total RNA
Extraction protocol RNA_seq: Primary hepatocytes: Cells were lysed in TRIzol-RNA lysis reagent (ThermoFisher). Liver tissue: Approximately 5 mg of liver tissues were homogenized using Ultra-Thorax in TRIzol-RNA lysis reagent. RNA was purified using EconoSpin columns (Epoc Life) according to manufacturer’s instructions.
ChIP-seq: Chromatin was prepared from 100-150mg formaldehyde crosslinked liver tissu. Crosslinked sonicated chromatin was IP'ed antibody and protein A/G agarose beads. DNA was extracted by phenol/choloroform.
Libraries from ChIP'ed DNA were constructed using the NEBNext Library prep kit according to manufacturer´s (NEB) instructions. Total RNA (1000 ng) was prepared for sequencing using ribosome depletion followed by random primed cDNA synthesis or polydT-mediated cDNA synthesis according to the manufacturer´s (Illumina) instructions. Subsequent library preparation was performed using the NEBNext RNA library preparation kit for Illumina.
 
Library strategy RNA-Seq
Library source transcriptomic
Library selection cDNA
Instrument model Illumina NovaSeq 6000
 
Description 8033_S84
Sample_8033
processed data file: RNAseq_RAW_CT_GFP_CRE.txt
Data processing RNA-seq and ChIP-seq sequencing data was aligned to mm10 or rn5 using STAR
RNAseq data was quantified by HOMER and normalised using DESeq2
GR ChIPseq peaks and bedgraph files were generated using HOMER
Genome_build: mm10 or rn5
Supplementary_files_format_and_content: Bedgraphs, peak bed file and text files with DESeq2 normalised RNAseq data
 
Submission date May 03, 2021
Last update date Sep 08, 2021
Contact name Lars Grøntved
E-mail(s) larsgr@bmb.sdu.dk
Phone +45 24 60 14 06
Organization name University of Southern Denmark
Department Biochemistry and Molecular Biology
Street address Campusvej 55
City Odense
ZIP/Postal code 5230
Country Denmark
 
Platform ID GPL24247
Series (1)
GSE173723 Impaired GR expression in liver disrupts feeding induced gene expression, glucose uptake and glycogen storage
Relations
BioSample SAMN18977467
SRA SRX10754420

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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