GSM7747218: HPV-mice, Replicate 2,  Blood; Mus musculus; RNA-Seq
Bone marrow single cell suspensions were generated from femurs and tibiae of mice. Cells were flushed by centrifuging the bones cut at the knee joint. The isolate was filtered through a 40 ?m cell strainer.  Single cell suspensions of spleens were generated by mashing the spleen through a 40 ?m cell strainer. Tumors were harvested and minced using a scalpel and digested for 30 minutes using collagenase A (0.33 U/mL, Roche), dispase (0.85 U/mL, Roche), and DNAse I (144 U/mL, Roche) in RPMI medium with intermittent shaking at 37°C. Afterward, tumor single cell suspensions were passed through a 70 ?m cell strainer. Blood was collected by intracardiac punction into EDTA-coated tubes. Red blood cells were lyzed with BD Pharm Lyse Lysing buffer (BD Biosciences, ref. 555899) according to the manufacturer?s instructions. Cell suspensions were further used for single cell RNA sequencing. Single-cell gene expression libraries were prepared following the manufacturer?s protocol with the 10x Genomics Next GEM Single Cell 5? Kit v2, and then sequenced on an Illumina HiSeq4000 platform.