RNA-Seq of liver of HFHC+AMEVmod1
RNA from liver samples (approximately 35 mg) was extracted using the RNeasy mini kit (Cat  #74104, Qiagen) following manufacturers instructions. To minimize technical variability, all liver samples were processed in parallel on the same day, thereby avoiding potential batch effects. Total RNA was quantified with NanoDrop (Thermo Fisher Scientific), and quality control (QC) was performed using an Agilent Bioanalyzer system (Agilent Technologies). The RNA libraries were sequenced on the Illumina NovaseqTM X Plus platform to generate the raw sequence reads. Raw sequencing reads were preprocessed with Cutadapt (v1.9.3) to remove adapter sequences and low-quality bases (Phred score <20), followed by quality assessment with FastQC to ensure data integrity. Clean reads were aligned to the mouse reference genome (GRCm39) using HISAT2 (v2.0.4) with default parameters. Transcriptome assembly and quantification were performed using StringTie (v2.2.1), which reconstructs both known and novel transcripts and accounts for alternative splicing events. Gene- and transcript-level expression values were calculated as FPKM (Fragments Per Kilobase of transcript per Million mapped reads) to enable cross-sample normalization and comparability.