Genome wide CRISPR screen for Venetoclax plus Ruxolitinib combination resistance in OCI-AML2 cells using Y. Kosuke library

30 million OCI-AML2 cells were infected with CRISPR/Cas9 library, selected for puromycin resistance for integration events and exposed DMSO, venetoclax (0.5 uM) or venetoclax (0.1 uM) plus ruxolitinib (1 uM).  Cells were collected at day 14 and 21 , DNA was extracted and sgRNA barcodes were amplified. PCR library was deep sequenced using Novaseq 6000 high throughput Illumina platform. Performed comparisons of sgRNA read counts between DMSO and Venetoclax as well as Venetoclax plus Ruxolitinib treated cells at corresponding time points.

三千万个OCI-AML2细胞经CRISPR/Cas9文库（CRISPR/Cas9 library）感染后，通过嘌呤霉素抗性筛选富集发生了基因整合的细胞，随后将细胞分为三组，分别用二甲基亚砜（DMSO）、0.5μM维奈克拉（venetoclax）以及0.1μM维奈克拉联合1μM芦可替尼（ruxolitinib）进行处理。分别在第14天和第21天收集细胞，提取基因组DNA并扩增sgRNA条形码序列，采用Illumina NovaSeq 6000高通量测序平台对构建的PCR文库进行深度测序。最后在对应时间点，比较二甲基亚砜组、维奈克拉单药组与维奈克拉联合芦可替尼组细胞的sgRNA测序读段计数差异。