title: GSM8657342: No treatment Day 0; Homo sapiens; OTHER
library_construction_protocol: The cells were infected at a multiplicity of infection of 0.3 to ensure that most cells took up only one stable short guide RNA (sgRNA). Puromycin (10 µg/ml) was added to the cells 72 h after infection and maintained in the cultures for 3 days. After effective selection for puromycin resistance, we equally divided these HT-29 cells into four groups: the DMSO (day 0), DMSO (day 14), LGX818 (1 μM/ml) treated for 14 days, or LGX818 (100 nM/ml) + cetuximab (100 ng/ml) treated for 14 days groups. After drug selection, the cells were harvested, and genomic DNA was isolated using a DNA extraction kit (Omega, D3396). The PCR products containing gRNA sequences that were integrated into the genomes of the cells were sequenced by parallel sequencing using a HiSeq 2000 (Illumina). Resistant genes were identified from sgRNA screen sequencing results using MAGeCK MLE analysis. MAGeCK MLE calculates a “beta score” for each targeted gene to measure the degree of selection upon gene perturbation, similar to the “log fold-change” measurement in differential expression analysis. A positive beta score indicates that the gene is a positively screened gene, whereas a negative value indicates a negatively screened gene.