GSM7747551: ABE-RA2.0_rep2_S32; Homo sapiens; OTHER
Transfected cells were incubated for an additional 3 days before harvesting. Genomic DNA was extracted using a Quick-DNA Miniprep Plus Kit (Zymo, D4069) following the manufacture's protocol. HEK293T cells were seeded into two 6 cm tissue culture dishes at a density of 1 x 106 cells per dish, and infected 18 h post-seeding with 2.5 mL lentiviral library at low multiplicity of infection (MOI) using 10 μg/mL polybrene. Cells were trypsinized 24 h post-infection, transferred into two 10 cm tissue culture dishes, and grown in DMEM medium supplemented with 10% FBS and 2 μg/mL puromycin. Cells were cultured and selected with 2 μg/mL puromycin for an additional 14 days. To maintain the diversity of the paired sgRNA-target library, cells were split and passaged at a minimum density of 1 x 107 cells per passage.