sgRNA targeted the exon 3 of PUMA gene, which contains sequence code for BH3 domain of PUMA. Two crRNAs introduced into lentiviral vectors (pLentiCRISPR-E, Addgene #78852) which contains eSpCas9 and puromycin cassette.

Guide1 DNA (forward, 5’-CACC GGCGGGCGGTCCCACCCAGG-3’; reverse, 5’-AAAC CCTGGGTGGGACCGCCCGCC-3’) and Guide 2 DNA (forward, 5’-CACC GCCGCTCGTACTGTGCGTTG-3’; reverse, 5’-AAAC CAACGCACAGTACGAGCGGC-3’) were annealed and ligated to the restriction enzyme-cut plasmid by T4 ligase. Stbl3 strain (Invitrogen C7373-03) was transformed by the guides-containing plasmids. LB-amp plates were streaked and incubated on a shaker at 37 °C overnight. The bacterial colonies were selected and mixed up with LB (Terrific Broth) and 100 μg/mL ampicillin, and were incubated on a shaker at 37 °C overnight. Plasmids from different colonies were isolated and purified using QIAprep Spin Miniprep Kit (Qiagen). Plasmids were digested with BsmBI and BamHI in Cut Smart Buffer (New England BioLabs, Inc.) at 37 °C for 1 h and then analyzed by 1% agarose gel. Sequencing was performed by GENEWIZ (South Plainfield, New Jersey, NJ; Fig. S5 A–F).

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