In vivo, PD1 screen, rep2
For CRISPR screen analysis, genomic DNA from Hepa1-6 cells (In vitro) or 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).
SUB16056251
Clone4_PD2