Longitudinal transcriptomic profiles of the small intestine and colon across the lifespan in a neonatal overfeeding mouse model

We employed an established neonatal overfeeding model by reducing litter size, assigning litters to either 4 pups per dam (Overfed group) or 8 pups per dam (Normal-fed control group). To longitudinally bridge early-life epithelial programming with later-life aging phenotypes at the transcriptomic level, small intestine and colon tissues were harvested from independent cohorts of mice at 3 weeks, 6 months, and 28 months of age. Total RNA was isolated from all aforementioned tissues using TRIzol Reagent followed by DNase I treatment (Takara) to remove genomic DNA contamination. RNA integrity and purity were verified using an Agilent 5300 Bioanalyzer (Agilent Technologies), with all samples exhibiting RIN values > 8.0. Library preparation was performed by Shanghai Majorbio Bio-pharm Biotechnology Co., Ltd using the Illumina Stranded mRNA Prep, Ligation protocol, followed by 150 bp paired-end sequencing on an Illumina NovaSeq 6000 platform.