Small RNA sequencing of lipoproteins (HDL) obtained from germ-free mice and facility matched specific-pathogen free mice.

Plasma was harvested from two cohorts of facility-matched germ free (GF) and specific-pathogen free (SPF) mice at the National Gnotobiotic Rodent Resource Center (NGRRC; North Carolina, USA). Plasma was then fractionated by size-exclusion chromatography using three tandem Superdex200 increase columns (Cytiva). Fractions corresponding with HDL were then pooled and concentrated prior to RNA isolation. Small RNA libraries were generated from total RNA using NextFlex V3 Small RNA Seq-kit (Perkin Elmer) according to manufacturer’s instructions. Equimolar amounts of each library were then pooled and sequenced on the NextSeq500 platform (Illumina). Individual libraries were then demultiplexed and analyzed with the TIGER analytical pipeline. GF (Case) vs SPF (Control)