GSM7502511: sgMIif2; Mus musculus; RNA-Seq
Three different tumors were dissociated to single-cell suspensions using collagenase I (2 mg/ml), collagenase IV (2 mg/ml), DNase I (1mg/ml) and a tissue dissociator. Cells filtered with a 40 μm filter were incubated with Red Blood Cell Lysis Buffer to remove red cells. Death cells were removed by Death Cell Removal Kit. Cells were resuspended in PBS and cell number was counted. Equal cells were collected for subsequent staining. Cells were then incubated with 20% FBS in PBS with 25 μg/mL of anti-mouse CD16/CD32 antibody  at 4°C for 10 minutes to minimize background antibody binding. Then cells were stained with FITC-anti-mouse CD45 at room temperature for 20 minutes, followed by PI staining for 5 minutes on ice. Cells after staining were sorted by the BD FACSAria II sorting system to harvest the FITC (CD45)-positive and PI-negative populations. Cells recovered were subjected to the 10X Genomics standard protocol for scRNA-seq. RNA libraries were prepared for sequencing using standard protocols