GSM6589264: replicate_7_AP1159_coT2; Mus musculus; RNA-Seq
Adjacent mucosa were dissociated by 9mM EDTA for 1h, and then followed by TrypLE™ Express Enzyme (1X) (ThermoFisher Scientific) incubation (37°C, 5min) to dissociate into single cells. Colon tumor cells were dissociated using the Miltenyi Biotec MACS Tumor Dissociation Kit (Miltenyi Biotec) according to manufacturer instructions. Colon tumors and adjacent mucosa tissues were collected and processed on the same day for scRNA-seq. Single cell Gene Expression Library was prepared according to Chromium Single Cell Gene Expression 3v3.1 kit (10x Genomics). In Brief, single cells, reverse transcription (RT) reagents, Gel Beads containing barcoded oligonucleotides, and oil were loaded on a Chromium controller (10x Genomics) to generate single cell GEMS (Gel Beads-In-Emulsions) where full length cDNA was synthesized and barcoded for each single cell. Subsequently the GEMS are broken and cDNA from each single cell are pooled. Following cleanup using Dynabeads MyOne Silane Beads, cDNA is amplified by PCR.  The amplified product is fragmented to optimal size before end-repair, A-tailing, and adaptor ligation. Final library was generated by amplification.