title: GSM8545048: Non-targeting plasmid library; synthetic construct; OTHER
library_construction_protocol: Plasmid libraries were purified using the Qiagen midi plus kit. A549 cells were subject to hybridization chain reaction (HCR) Flow-FISH as in. (https://pubmed.ncbi.nlm.nih.gov/34326544/). Briefly, cells were fixed in 4% formaldehyde, permeabilized using 70% ethanol, FISH targeting PER1 were hybridized, and then the signal was amplified using the B3 amplifier (molecular instruments). Cells were washed and then top and bottom 12% PER1 cells were sorted based on PER1 expression (FITC). Genomic DNA was extracted from these cells using the FFPE miniprep kit (Zymo). Genomic DNA was also extracted from a bulk unsorted cell population using a Puregene kit (Qiagen) crRNA arrays were amplified from genomic or plasmid DNA using Q5 Hotstart 2x master mix. Primers contained Illumina adapter sequences and indexes