title: Illumina HiSeq 2500 sequencing; Transcriptome profiling of liver samples of C/EBPß ∆uORF mice
design: Transcriptome profiling of liver samples of C/EBPß ∆uORF mice
library_name: oKI6_s
library_construction_protocol: Knock-in in the Cebpb locus that replaces the wt Cebpb-mRNA sequence for a mutated version that contains a mutation in the AUG-initiation codon of the cis-regulatory upstream open reading frame (uORF), abrogating uORF function. The exact mutation is wt AGCATGCCT -> delta-uORF AGAATTCCT (introducing EcoRI site). This mutation results in deficiency of uORF-dependent translation into the C/EBPbeta-LIP protein isoform, leaving translation into the C/EBPbeta-LAP protein isoforms unaffected. Liver tissue from young (5 months) and old (20 months) wt and C/EBPβΔuORF mice (from six individuals per group) was homogenized on ice with a motor driven pellet pestle (Kontes) in the presence of QIAzol reagent (Qiagen) and total RNA was isolated as described in Zidek et al., 2015. RNA was isolated from tissue samples using the RNeasy® Lipid Tissue Mini Kit (QIAGEN). After incubation for 30 min at 37°C with RQ1 RNase‐Free DNase (Promega), the RNA was further purified with the RNeasy® Plus Mini Kit (QIAGEN) according to the manufacture's protocol starting at step 4. Preparation of the sequencing libraries was performed using the TruSeq Sample Preparation V2 Kit (Illumina) according to the manufacturer's instructions.
attributes.tag: Experimental Factor: genotype
attributes.value: C/EBP uORF expression
attributes.tag: Experimental Factor: age
attributes.value: 19