title: GSM7913146: UPAR- Male; Mus musculus; RNA-Seq
library_construction_protocol: Small intestine was removed, washed with cold PBS, opened laterally and cut into 3-5mm fragments. Pieces were washed multiple times with ice cold PBS until clean, washed 2-3 with PBS+EDTA (7.5mM), and incubated with mild agitation for 30 minutes at 4C. Crypts were then mechanically separated from the connective tissue by shaking, and filtered through a 70-μm mesh into a 50mL conical tube to remove villus material and tissue fragments. For Epcam+ cell isolation, the crypt suspensions were dissociated to individual cells with TrypLE Express (Thermo Fisher Scientific, 12604039) and stained for flow cytometry. Epithelial cells were isolated as SYTOX-, CD45- Epcam+ with a BD FACS Aria II SORP cell sorter into supplemented crypt culture medium for culture. uPAR+ and uPAR- populations were isolated as DAPI-, uPAR+/- with a SONY cell sorter(SH800S). Single cell libraries were prepared using a 10X Genomics Chromium Controller (cat Chromium Next GEM Single Cell 3' Gene Expression kit (cat #1000268) according to the manufacturer's instructions. Cell suspensions were adjusted to target a yield of 8,000 cells per sample. Single cell 3' digital gene expression with cell barcodes and unique molecular identifiers (UMIs).