sgRNA nucleotides were cloned into pLKO5-sgRNA-EFS-GFP (Addgene) as described in (48) with Bsm BI digestion. pLKO5-sgRNA-EFS-GFP and pHR-SFFV-KRAB-dCas9-P2A-mCherry (Addgene) were used for lentiviral supernatant production by transfection of HEK cells.

shRNA-mir oligonucleotides were cloned into the pMSCV-LTRmiR30-SV40-GFP vector using Eco RI restriction digest. Retroviral supernatant was produced by transfection of Phoenix-ECO cells.

Expanded, tertiary Lp30 cell vials were thawed and cultured in x-vivo complete media. One day after thawing, cells were transduced twice on two consecutive days by 50-min retro- or lentivirus spinoculation (2000g, 32°C) on RetroNectin (Takara Bio)–treated and 2% BSA (STEMCELL Technologies)–blocked wells, and cells were plated subsequently at 3 × 105 cells per milliliter of X-VIVOTM complete media. For analysis after transduction, cells were resuspended in phosphate-buffered saline (PBS) + 3% fetal calf serum.

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