GSM9229090: iWAT, PM2.5, 1; Mus musculus; RNA-Seq
Focal areas of solid and GG components were identified by tissue appearance and compute tomography (CT) at the same time. Different components were macrodissected and stored at 4 ℃ GEXSCOPETM Tissue Preservation Solution. Then, we washed samples for 3 times with Hanks Balanced Salt Solution (HBSS) and minced them into 1–2 mm-thick pieces, 2 ml GEXSCOPETM Tissue Dissociation Solution (Singleron Biotechnologies) was added to each sample and allowed for digestion at 37 ℃ for 15 minutes to dissociate the tissue into single cells. Then, we filtrate the samples using 40-μm sterile strainers, followed by centrifuging at 1,000 rpm for 5 minutes. After the centrifugation, the precipitation was resuspended in 1 ml phosphate-buffered saline (PBS; HyClone, Marlborough, United States). Subsequently, 2 ml GEXSCOPETM Red Blood Cell Lysis Buﬀer (Singleron Biotechnologies) was added into the cell suspension and incubated at 25°C for 10 minutes to remove red blood cells. After that, the mixture was centrifuged at 500 × g for 5 minutes, followed by re-suspending the precipitation with PBS. We used GEXSCOPETM Single Cell RNA Library Kit Tissue (Singleron Biotechnologies) to barcode single cells, capture mRNA from isolated single cells, and generate cDNA libraries for scRNA-seq. Then, individual libraries were diluted to 4 ng/μl, and pooled for sequencing on a Illumina NovaSeq 6000 with 150 bp paired-end reads.