GSM8182184: young mice; Mus; RNA-Seq
Tissues were washed three times with PBS, cut into blocks of 2 mm and placed on DMEM-1640 medium (Gibco, Gaithersburg MD, USA).  Using buffer containing collagenase IV (1 mg/ml; Thermo Fisher), pronase E (1 mg/ml), and DNase I (50 U/ml), the tissues were digested at 37°C for 30 min.  Cells were filtered through a 70um cell strainer and centrifuged at 400×g for 5 minutes.  Take a small amount of single-cell suspension, add 0.4% trypan blue staining solution of the same volume, count the cells with Countess II Automated Cell Counter, and adjust the concentration of living cells to the ideal range (1000 to 2000 cells/μl). The gel beads containing barcode information combine with a mixture of cells and enzymes,then enter the reservoir to be separated by oil to form GEMs (GelBeads-In-Emulsions).  After that, the gel beads dissolved and released the capture sequence containing the Barcode sequence, reverse transcribed the cDNA fragment, and labeled the sample.  Break up the gel beads and oil droplets then use cDNA as a template for PCR amplification.  All the products of GEMs are mixed to construct a standard sequencing library.  The cDNA was cut into 200~300bp fragments, then the second-generation sequencing library was constructed.  Finally, the DNA library was amplified by PCR.  The high-throughput sequencing of the library was carried out by using the PE150 sequencing mode of the Illumina sequencing platform.  Quality control of sequencing was performed using cellranger.