title: GSM7809269: Brown adipose tissue Control; Mus musculus; RNA-Seq
library_name: GSM7809269
library_construction_protocol: Fresh Brown fat depots nuclei were collected from 10-week-old JunB FKO and control mice. Tissue samples were minced into 1 mm3, homogenized in Nuclei EZ Lysis Buffer (NUC101, Sigma-Aldrich) with pre-chilled Dounce homogenizer. The homogenate was filtered using a 40-μm cell strainer (VWR#76327-098). After centrifugation, the pellet was washed, re-suspended, and stained with DAPI, the single and health nuclei were isolated using sorting. Reconstitute nuclei with concentration of 700-1,200 nuclei/μl was used for the sequencing. Single-nuclei RNA libraries were prepared according to the Chromium™ Single Cell 3' Reagent Kits v2 User Guide (10x Genomics). Approximately 10,000 cells were loaded on a Chromium single Cell Controller Instrument (10x Genomics) to generate single cell gel beads in emulsion (GEMs). The barcoded sequencing libraries were constructed using the Chromium Single-Cell 3'Library Kit (10x Genomics) for enzymatic fragmentation, end-repair, A-tailing, adaptor ligation, ligation cleanup, sample index PCR, and PCR cleanup. Libraries were sequenced with NovaSeq System with a depth of 50,000to 100,000 reads per targeted cell.