Single-cell DNA sequencing analysis of ecDNA copy-number dynamics during drug adaptation
Single-cell DNA sequencing analysis of ecDNA copy-number dynamics during drug adaptation
COLO320DM, COLO320HSR, and RPE1 cells were cultured in Dulbecco's Modified Eagle's Medium (DMEM) low glucose supplemented with 10% FBS. H2170, SNU-16, and SK-BR-3-Luc cells were cultured in Roswell Park Memorial Institute (RPMI) 1640 Medium supplemented with 10% FBS. PC-3 cells were cultured in Kaighn's modification (F12K) of Ham's F12 medium supplemented with 10% FBS. All cells were incubated at 37℃, 20% O2 and 5% CO2. Cells were processed using the Mission Bio Tapestri platform according to the manufacturer's instructions. Within droplets, targeted PCR amplification was performed to generate single-cell DNA libraries. The library was prepared according to the manufacturer's instructions. Briefly, following targeted PCR, the emulsion was disrupted, the reaction was enzymically cleaned up, and the product were purified with AMPure XP beads. To add indexes, 10 cycles second PCR was performed, followed by purification with AMPure XP beads. Library quality was assessed by Quantus and Tapestation 4200 with D5000 Screen Tape. 