title: GSM9270220: KPC1199, vehicle_2; Mus musculus; RNA-Seq
library_construction_protocol: Under sterile conditions, freshly collected KPC1199 mouse tumor tissues were washed twice with pre-chilled RPMI 1640 medium supplemented with 0.04% BSA. The tissues were thoroughly minced into small pieces using surgical scissors and subsequently placed in a freshly prepared enzymatic digestion solution. Digestion was carried out for 30-60 minutes in a 37°C incubator, with the mixture being inverted every 5-10 minutes to ensure uniform dissociation.The digestion solution consisted of RPMI 1640 (Corning, cat. no. 10-040-CVR), 0.04% BSA (MACS, cat. no. 1000076), and 0.2% Collagenase II (Gibco, cat. no. 17101015).Following digestion, the resulting cell suspension was filtered once or twice through a 40 μm cell strainer (BD Falcon, cat. no. 352340) and centrifuged at 300 x g for 5 minutes at 4°C. The cell pellet was resuspended in a suitable volume of medium, and an equal volume of Red Blood Cell Lysis Buffer (Miltenyi, cat. no. 130-094-183) was added. The mixture was incubated at 4°C for 10 minutes. The cell suspension was then centrifuged at 300 x g for 5 minutes, and the supernatant was discarded. The pellet was washed once more with medium by centrifuging at 300 x g for 5 minutes and discarding the supernatant. Finally, the cells were resuspended in 100 μl of RPMI 1640 medium (Corning, cat. no. 10-040-CVR) containing 0.04% BSA. The concentration and viability of the single-cell suspension were determined using a Luna-FL™ Automated Fluorescence Cell Counter (Logos Biosystems, Korea) or by manual counting with trypan blue staining. KPC1199 tumors were processed to single cell suspension and the directly used for library extraction. Library was constructed using Chromium Next GEM Single Cell 3' Reagent Kits version3.1 (10x Genomics)