In vivo, OT1 screen, rep6
For CRISPR screen analysis, genomic DNA from tumor tissues (In vivo) were extracted using Allprep DNA Mini Kit (QIAGEN). Then the region containing sgRNA cassette was amplified using primers specific to the lentiGuide-puro vector. PCR was performed using Q5 High-Fidelity DNA Polymerase (NEB), 200 g of gDNA was used per sample (2 g of gDNA per reaction). The PCR products were pooled and gel purified from a 2% agarose gel using the MinElute Gel Extraction kit (QIAGEN). The pooled products with 10%-20% PhiX were sequenced on HiSeq 2500 system (Illumina).