GSM8547349: Pancreatic ductal adenocarcinoma PDO, TRACER Layer 2; Homo sapiens; RNA-Seq
After 24h of culture in the rolled configuration, TRACER constructs were disassembled and sectioned into individual layers. The indiviudal layers were then incubated in a solution of HBSS supplemented trypsin 0.5x for 30 mins on an orbital shaker at 900 rpm and 37C. Samples were agitated by repeated pipetting to dislodge the organoid cells and the digestion reaction was neutralized wtih Advanced DMEM/F-12 media supplemented with 10% FBS. Cells were collected by centrifugation. The library was prepared in accordance with manufaturer's instructions (Single Cell 5' v2 Reagent Kits, 10x genomics). Briefly, cells were washed and assessed for viability, then loaded onto a 10X single cell chip K. cDNA was recovered using a 10X recovery agent and cleaned using Silane DynaBead mix as outlined in the user guide. Purified cDNA was amplified and cleaned using SPRI select beads. 5' v2 cDNA libraries were then prepared as outlined in the Single Cell 5'v2 Reagent kid user guide.