description: Protocols: Murine tissues for protein or RNA extraction were frozen at the time of collection and stored at -80. All data are from male mice. Insig1 KO mice were generated on a C57Bl6/J background and compared to wild-type and heterozygous littermates. Mice were maintained in a room with controlled temperature (21°C; 12 hr light/dark cycle) with free access to food and water; the pathogen-free facilities were compliant with the FELASA Guidelines. The UK Home Office and the USC Bioethics Committee of the Universities of Cambridge and Newcastle approved all animal procedures. Mice were fed on a chow diet (Safe Diets, Code ds-105) until they were enrolled in specific procedures. WDSW (Western Diet + Sugar Water) and LFD (Low Fat Diet) Animals; At the age of 8 weeks, mice were fed either Low Fat Control (Teklad, TD08485) or Western Diet (Teklad, TD88137) plus drinking water supplemented with fructose and glucose (SW, 23.1 g/L d-fructose +18.9 g/L d-glucose) starting from week 8; total duration of the experimental challenge was 12 weeks. Fat/lean mass was calculated by time-domain nuclear magnetic resonance (TD-NMR) by using a minispec Live Mice Analyzer LF50 (Bruker) the day of the culling. Serum and tissue collection were performed in fed condition the day of the culling. Murine RNA was isolated using miRNAeasy Mini Kits (Qiagen), according to the manufacturer's instructions. All reagents, plastic ware, and supplies used were nuclease-free, sterile, and of molecular biology grade. RNA purity (A260/A280>1.80) and concentration were determined using the Nanodrop spectrophotometer (Thermo Fisher Scientific, Delaware USA). RNA integrity was studied using the 2100 Bioanalyzer (Agilent) and RNA 6000 Nano Kits (Agilent, Santa Clara, California, USA). RNA Integrity Number (RIN) of 8 was considered the lowest cut-off for RNA sequencing. RNA from tissues (1 ug RNA) were used to generate barcoded sequencing libraries using Illumina TruSeq® Stranded mRNA Library Preparation Kit (Illumina) following manufacturer's instructions. Library preparation was performed by the Genomics and Transcriptomic Core at the Institute of Metabolic Science.
attributes.tag: ENA first public
attributes.value: 2021-04-15
attributes.tag: ENA last update
attributes.value: 2020-12-09
attributes.tag: External Id
attributes.value: SAMEA7699102
attributes.tag: INSDC center alias
attributes.value: Co-author
attributes.tag: INSDC center name
attributes.value: Co-author
attributes.tag: INSDC first public
attributes.value: 2021-04-15T00:32:17Z
attributes.tag: INSDC last update
attributes.value: 2020-12-09T12:53:31Z
attributes.tag: INSDC status
attributes.value: public
attributes.tag: Submitter Id
attributes.value: E-MTAB-9864:Sample 29
attributes.tag: age
attributes.value: 8
attributes.tag: broker name
attributes.value: ArrayExpress
attributes.tag: common name
attributes.value: house mouse
attributes.tag: developmental stage
attributes.value: adult
attributes.tag: diet
attributes.value: low fat diet
attributes.tag: disease
attributes.value: normal
attributes.tag: duration of experiment
attributes.value: 12
attributes.tag: genotype
attributes.value: wild type genotype
attributes.tag: individual
attributes.value: D708_D506_LFD
attributes.tag: organism part
attributes.value: liver
attributes.tag: sample name
attributes.value: E-MTAB-9864:Sample 29
attributes.tag: sex
attributes.value: male
attributes.tag: strain
attributes.value: C57Bl6/J