Expression profiling by high throughput sequencing
Summary
We identify sestrins, a family of stress-inducible metabolic regulators, as protective factors against muscle wasting. Sestrin expression decreases during inactivity and its genetic deficiency exacerbates muscle wasting; conversely, sestrin overexpression suffices to prevent atrophy.
Overall design
RNAseq analysis: total RNA from tibialis anteror TA muscles was extracted using a protocol combining QIAzol Lysis Reagent and RNAeasy minikit columns (Qiagen) following the manufacturer’s instructions. RNAseq services were provided by the CNIC Genomics Unit, including quality control tests of total RNA using Agilent Bioanalyzer and Nanodrop. spectrophotometry. cDNA library preparation and amplification were performed from 200 ng total RNA using NEBNext Ultra RNA Library Prep Kit for Illumina. RNAseq analysis was performed with 3-4 samples per condition, using Illumina Hiseq 2500. RNAseq analysis was done in two sets. Sets are indicated in the sample and file names (Set1 and Set2).
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