GSM8242227: Inguinal white adipose tissue, PTRF knockout adipose tissue from KNOCKOUT1 mice, snRNAseq; Mus musculus; RNA-Seq
snRNA-seq samples was extracted from iWAT of 6-month-old mice(n = 2). iWAT were isolated immediately and stored in the sCelLiveTM Tissue Preservation Solution (Singleron) on ice after the surgery within 30 mins. The samples were washed in pre-cooled PBSE for three times, minced into small pieces, and then dissociated into nuclear suspensions using GEXSCOPE® Nucleus Separation Solution (Singleron Biotechnologies, Nanjing, China) refer to the manufacturer's product manual. The mixture was then centrifuged at 300 × g 4 ℃ for 5 mins to remove supernatant and resuspended in PBSE to 10^6 nuclei per 400μl. The isolated nuclei suspension was collected and filtered through a 40-micron sterile strainer. Finally, cell number and cell viability were measured by trypan blue staining microscopically. Nuclei were counted in a Luna cell counter and a nuclear density of 3-4 × 10^5 nuclei/mL were loaded onto a microfluidic chip (GEXSCOPE® Single NucleusRNA-seq Kit, Singleron Biotechnologies) and snRNA-seq libraries were constructed according to the manufacturer's instructions (Singleron Biotechnologies). Barcoding Beads are subsequently collected from the microwell chip, followed by reverse transcription of the mRNA captured by the Barcoding Beads and to obtain cDNA, and PCR amplification. The amplified cDNA is then fragmented and ligated with sequencing adapters. Individual libraries were sequenced on Illumina novaseq 6000 with 150 bp paired end reads.